Regulation of membrane lipid synthesis in Escherichia coli. Accumulation of free fatty acids of abnormal length during inhibition of phospholipid synthesis. Glycerol starvation of an Escherichia coli glycerol auxotroph results in a specific inhibition of membrane phospholipid synthesis. Mindich ((1972) J. Bacteriol. 110, 96-102) observed only a trace accumulation of free fatty acid following glycerol deprivation. We have repeated these experiments using glycerol auxotrophs which also possess a lesion in beta oxidation. This defect was introduced in order to control fatty acid degradation. In contrast to the previous results, we find free fatty acid does accumulate during glycerol starvation. Similar results were found using beta oxidation-defective (fadE-) derivatives of both gpsA and plsB glycerol auxotrophs. Upon glycerol starvation of a plsB- fadE- strain, phospholipid synthesis is 90 percent inhibited. Following a lag of 20 to 40 min, free fatty acid synthesis begins and proceeds at a rate that steadily increases until the rate of fatty acid synthesis is equal to that found in glycerol-supplemented cultures. The accumulation of free fatty acid is the result of de novo synthesis.  The average chain length of the fatty acid in the unesterified fraction is abnormally long. Two 20-carbon fatty acids, cis-13-eicosenoic acid and arachidic acid, are found in this frction. Furthermore, a greatly increased level of stearic acid and a small amount of a C-22 (behenic) acid are found in the free fatty acid fraction. These data indicate that acyl transfer into phospholipid is a major determinant of phospholipid acyl moiety chain length. Other experiments have shown that the free fatty acid fraction in glycerol-starved cells is metabolically active. This fraction turns over despite the defective beta oxidation system. Restoration of glycerol to starved cells allows the incorporation of the unesterified fatty acids into phospholipid.


Ten proteins required for conversion of phiX174 single-stranded DNA to duplex form in vitro. Resolution and reconstitution. Protein requirements for conversion of phiX174 single-stranded DNA to a double-stranded replicative form with a small gap (RF II) have been determined by resolution and reconstitution of the multienzyme system from extracts of gently lysed Escherichia coli. Assays depended on: (a) complementation of extracts of thermosensitive mutants and (b) fractionation of extracts of wild type cells to divide essential components into groups, each of which was further resolved. These procedures have yielded eight proteins: dnaB protein, dnaC protein, proteins i and n (two novel proteins without a defined genetic locus), dnaG protein, DNA polymerase III holoenzyme (polymerase III and copolymerase III), and DNA unwinding protein; purification procedures for the first four are presented here. (Closure of RF 22 requires as with phage M13, DNA polymerase I and ligase.)


dnaG gene product, a rifampicin-resistant RNA polymerase, initiates the conversion of a single-stranded coliphage DNA to its duplex replicative form. The protein responsible for the initiation of conversion of single-stranded phage G4 DNA to the duplex replicative form has been purified approximately 3000-fold and identified with Escherichia coli dnaG gene product. The protein is a rifampicin-resistant RNA polymerase of approximately 64,000 daltons. It catalyzes the incorporation of the four ribonucleoside triphosphates into an oligoribonucleotide, using as template the single-stranded DNA coated with the DNA unwinding protein of E. coli. An RNA transcript of a unique region of the chromosome can serve as a primer by covalent extension by DNA polymerase III holoenzyme to form a nearly full-length linear complementary strand. A similar role for the dnaG protein in the initiation of nascent (Okazaki) fragments in replication of the host chromosome is discussed.


Myelodysplastics--fate of those followed for twenty years or more. Of a total of 143 myelodysplastic patients treated between 1928 and 1951, there were sixty-three patients with severe myelodysplasia whose records allowed long-term review. At the time of writing twenty-nine were alive and were twenty to forty-three years old. All were walking in the hospital while under an intensive physical therapy program. However only two of the nine with twelfth thoracic-second lumbar function were walking at final follow-up as adults, while nineteen of the twenty with function at the third lumbar level were doing so. The status of the hips did not correlate with the ability to walk. One-third of the survivors were self-supporting at the time of writing. About one-half had scoliosis and in one-third was greater than 20 degrees.


The use of flow microbluorimetry in the analysis of the phenotype expression of mouse histocompatibility antigens. Quantitation of the expression of cell surface antigens has hitherto been limited to analysis by either cytotoxicity tests or radioimmune assays (5, 15). We report here the use of a new methodology to analyze and quantitate the expression of mouse histocompabililty antigens (H-2 locus) in hybrid clones and parental cell types. The binding of fluorescein-tagged antibody is measured on a cell-to-cell basis in large viable cell populations using flow microfluorimetric techniques. These techniques have been used to measure hapten and immunoglobulin binding to lymphocyte populations (8, 9, 14). However, this is the first report in which these techniques have been used to examine the expression of the H-2 locus. The advantage of this approach is twofold: first, a large and statistically significant sample population may be analyzed one cell at a time, thus revealing the fine detail of heterogeneity in the expression of the cell surface markers within a population. Second, as has been demonstrated for analysis of specific components of the immune system, this method does permit fluorescence-activated sorting of cell types according to their different surface populations (8, 9, 14).


Fine structure of an organelle associated with the nucleus and cytoplasmic microtubules in the cellular slime mould Polysphondylium violaceum. Polysphondylium violaceum was grown in association with Escherichia coli. Vegetative amoebae and pseudoplasmodia were fixed under different conditions and processed for electron microscopy. An electron-opaque body (nucleus-associated body, NAB) lies in the cytoplasm near the tapered end of interphase nuclei. The NAB consists of a disk-shaped, multilayered core, approximately 200 nm in diameter and 150 nm thick, embedded in a granular matrix from which electron-opaque nodules protrude. The nodules are termination points of microtubules radiating from the NAB into the cytoplasm or running along the nucleus. On the average there are 16 nodules per NAB. One or two microtubules terminate in each nodule. Spindle pole bodies, arising by duplication of the NAB at the beginning of mitosis, are unstructured foci for spindle microtubules in mitotic cells. It is suggested that cytoplasmic microtubules do not determine cell shape, but they probably cause the tapering deformation of the nucleus. They may, furthermore, represent a storage form of subunits for utilization during the formation of the mitotic spindle. The nodules of the NAB are potential nucleation sites of cytoplasmic microtubules during interphase. Spindle pole bodies presumably acquire a microtubule organizing capability by integration of the decondensed nodules.


Chromatography of acid proteinases and chymotrypsin on a sorbent containing 2,4-dinitrophenyl residues. A sorbent obtained by treatment of cyanogen bromide-activated Sepharose 4B with mono-N-DNP-hexamethylenediamine has been shown to be effective in the affinity chromatography of pepsin, pepsinogen and acid proteinase from Aspergillus awamori. It is considered that 2,4-dinitrophenyl residues of the sorbent interact specifically with the hydrophobic zone of the enzyme, which may belong to the substrate binding site. The chromatography of chymotrypsin on the same sorbent supports this assumption.


Tolerance and resistance of myocardium to anoxia: experimental studies. We shall emphasize the notion of wide variability in the tolerance and resistance to anoxia of the myocardium, although the most important parameters are temperature and time. They can be modified widely by accompanying physical or biochemical conditions: --Cardiac distension and drugs inducing cardiac arrest; --Resuscitative perfusion; --The effect of some biochemical components is more likely explained by their possible action on the mitochondriae (glyoxilic acid-Isoproterenol). The study of the mitochondriae alone can explain the different behaviours of hearts subjected to anoxia in various situations.


Intraoesophageal rupture of a thoracic aortic aneurysm. The intraoesophageal rupture of a large thoracic aortic aneurysm is reported in a 49 year old man. He had been hypertensive for some years while the aneurysm increased in size. Although a graft was successfully inserted to repair the leak, infection from the oesophagus with candida albicans, subsequently led to secondary haemorrhage and death 17 days later. A plea is made for the earlier referral of patients with aneurysm prior to rupture, as the operative mortality rises markedly after rupture has occurred and in this case the situation was virtually irreparable.


LH and FSH response to gonadotropin-releasing hormone in anorexia nervosa: Effect of nutritional rehabilitation. The LH and FSH responses to the administration of synthetic gonadotropin-releasing hormone (GnRH) were assessed in 14 patients with anorexia nervosa before, during and following weight gain. In 8 patients studied prior to therapy when they were at 53-64% of idal body weight, the LH response to GnRH was strikingly impaired while the FSH response was equal to that observed in normally menstruating women. After return to 79-88% of ideal body weight the LH response was improved but remained less than in the normal women. In a second group of 6 patients who were studied 4-11 months after hospital discharge when they were 90-94% of ideal body weight, the LH response was always equal to or greater than that found in the normal women. The serum estradiol concentration was low in all of the patients and remained less than 33 pg/ml even in those who returned to more than 90% of ideal body weight. Treatment with 25 mug of ethinyl estradiol for 3 days prior to repeat administration of GnRH did not result in a statistically significant increase of the LH or FSH responses above those before treatment. These data suggest changes occur in both hypothalamic and pituitary regulation of gonadotropin secretion in the course of nutritional rehabilitation of patients with anorexia nervosa and that synthesis and secretion of LH and FSH are modulated independently at the level of the pituitary gland.


Effect of synthetic gonadotropin-releasing hormone (GnRH) in a patient with the "fertile eunuch" syndrome. A patient with isolated LH deficiency ("fertile eunuch" syndrome) was given synthetic gonadotropin releasing hormone (GnRH) by a single intravenous injection and serum levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH), testosterone and dihydrotestosterone were measured. A significant rise in both gonadotropins was found after GnRH and the increase was similar to that reported for normal men. No significant change was noted in the serum androgen levels. The patient was re-tested with another single intravenous injection after a 6-h infusion of GnRH two days later. The rise in serum Fsh and LH levels during the infusion was greater than with the initial bolus. A further increase in LH but not FSH concentration was noted when a 150 mug bolus was given at the end of the infusion. Acute (4 day) treatment with human chorionic gonadotropin (hCG) resulted in an increase in the serum testosterone and dihydrotestosterone levels to those observed in normal adult males. The "fertile eunuch" syndrome thus appears to be a hypothalamic disorder. It is consistent with the concept of more than one hypothalamic factor controlling gonadotropin secretion. Other possibilities would be deficient or defective production of a releasing factor affecting one or both gonadotropins. The lack of a definite response of serum androgens to GnRH would appear to be secondary to a decreased number of differentiated Leyding cells.


The role of estrogen in the modulation of pituitary sensitivity of LRF (luteinizing hormone-releasing factor) in men. The role of estradiol in modulating pituitary gonadotropin release in the human male was studied by evaluating the effects of clomiphene on the pituitary gonadotropin response to synthetic LRF (150 mug) in 6 eugonadal men. Administration of clomiphene (100 mg single daily dose time 5 days) induced a significant elevation of the basal levels of LH, FSH, estradiol and testosterone. However, the pituitary release of LH and FSH in response to LRF was markedly diminished by the clomiphene treatment. This finding suggests that in men, as in women, endogenous estradiol provides feedback regulation of gonadotropin output by the pituitary.


Pressure tension, and force of closure of the human lower esophageal sphincter and esophagus. The mechanical characteristics of the circular muscle of the human lower esophageal sphincter and esophagus were studied in subjects with competent and incompetent sphincters. Pressure-diameter curves were constructed by producing various degrees of circumferential stretch with pressure-measuring probes of increasing diameter. The circumferential membrane tension (force of closure) and the circumferential stress (muscle tension) of the circular muscle layer were also calculated from these data. The pressure-diameter curves of competent and incompetent sphincters were different in magnitude and shape. Incompetent sphincters had lower pressures at all diameters, with pressure gradually increasing with larger probe diameter. In contrast for competent sphincters the pressure was highest near closure, with an initial decline and then an increase in pressure with increasing probe diameter. Both shape and magnitude of pressure-diameter curves of competent and incompetent sphincters were interchangeable when manipulated by pharmacologic agents. Urecholine increased the pressures and changed the incompetent pressure-diameter curve to the levels of the competent sphincter; conversely. Pro-Banthine decreased pressures and changed the shape of the competent pressure-diameter curve to the levels of the incompetent sphincter. Force of closure and circular muscle tension curves of competent and incompetent sphincters were similar in shape but were higher at all diameters for competent sphincters. Force of closure and circular muscle tension increased with larger probe diameter. However, the diameter of optimal tension development was larger than the largest probe used and certainly far from closure. Fundoplication increased the magnitude and changed the shape of the incompetent pressure-diameter curve to one similar to a competent curve. This pressure change was associated with an increase in the force of closure, suggesting that fundopliation modified the length-force of closure characteristics of the incompetent spincter.


Serological diagnosis of congenital toxoplasmosis. A comparison of the indirect haemagglutination test (IHAT) with the dye test (DT) in 23 infants with congenital toxoplasmosis showed that in the early part of the first year of life the IHAT titres were nearly all lower than the DT titres. In six cases the IHAT was negative or the titre was so low that the cases could have been missed in routine screening. Later in the first year the IHAT titres became higher than the DT and remained so. The infants belonging to mothers with a high IHAT and therefore assumed to have become infected early in pregnancy had severe infections which included all the cases of hydrocephalus. Infants of mothers with low IHAT titres relative to the DT and therefore thought to have been infected later in pregnancy had generalized, mild or subclinical infection. With some difficulty a suitable commercial conjugated anti-IgM serum was found that gave positive results in immunofluorescent antibody tests for specific IgM antibody with all the infected infants and their mothers throughout the first year and negative results in uninfected infants and their mothers.


Effects of delayed culture on semiquantitative urinary bacteriology results. Urine specimens received in the laboratory were cultured at intervals of two hours on three successive occasions. During the intervening period the specimens were left at room temperature. After 24 hours' incubation counts were done, and it was observed that the significance of bacterial growth was not altered by delay in culture.


Effects of minocycline and tetracycline on the vaginal yeast flora. Two groups of women with non-specific genital infection were treated for two weeks with tetracycline 250 mg six-hourly and minocycline 100 mg twice daily respectively. Cultures for yeasts were performed before treatment and at the end of the first and second weeks. Before treatment yeasts, mostly Candida albicans, were recovered from 13% of the women. After one and two weeks' treatment yeasts were isolated from 22 and 29% of women treated with tetracycline, and from 19 and 29% of women treated with minocycline. It is concluded that despite inhibition of the growth of yeasts shown by minocycline in vitro, there is no evidence of any significant difference between the actions of tetracycline and minocycline on the vaginal yeast flora.


A study of immunoglobulin M antibody to measles, canine distemper, and rinderpest viruses in sera of patients with subacute sclerosing panencephalitis. Seven boys were studied who had the clinical features of subacute sclerosing panencephalitis (SSPE) and whose brain histology was consistent with SSPE. Measles antigen was detected in the seven brains by the direct fluorescent antibody method. Three out of the seven boys had in their sera measles specific immunoglobulin M (IgM) which was detected by the indirect fluorescent antibody method, and the cell receptors for it were acetone stable. A prozone effect was noted in the sera of two patients. The absorption of one patient's serum with Staphylococcus aureus to reduce the titre of immunoglobulin G (IgG) removed the prozone effect. Two of the boys who had high titres of measles specific IgM also had serum IgM which reacted with canine distemper virus antigen but the titres were eightfold lower. None of the boys had detectable rinderpest specific IgM in their sera.


Interaction in vitro between synovial cells and autologous lymphocytes and sera from arthritis patients. Synovial cells from patients with rheumatoid arthritis (RA) when grown in vitro in media supplemented with 20% fetal calf serum failed to show any difference in growth rate, life span, uptake of tritiated thymidine or cellular and nuclear characteristics when compared with synovial cells from patients with osteoarthritis or other joint diseases grown similarly in 20% serum enriched medium. There was also no evidence that lymphocytes and/or sera from RA patients were more cytotoxic to autologous synovial cells than sera and/or lymphocytes from OA patients. It is unlikely that antisynovial antibodies or lymphocytes from RA patients act as triggers for synovial damage.


Use of a computer for producing microbiological reports and for data storage and processing. A simple 'fail-safe' system is described that produces microbiology reports on a computer teleprinter and stores results on the magnetic tapes and discs of a computer for instant retrieval and epidemiological analysis. The system, which has been in operation for over a year, involves the use of a modified conventional NCR request form. The top portion, which is completed manually by the laboratory staff, is coded by writing numbers in hatched boxes. The data thus written on to the bottom portion are transcribed by punch operators on to paper tape and this data input is verified by double punching. The reports are normally produced automatically by the computer terminal telprinter, but in case of mechanical failure the manually completed request forms can be returned to wards and outpatient departments. The system permits a wide choice of options for epidemiological analysis, and six programmes are described, one of which produces a digest of  the overall percentage antibiotic sensitivities of organisms from various sites.


Study of adjuvant chemotherapy in osteogenic sarcoma. Twelve patients with osteosarcoma but without metastases were treated with adjuvant chemotherapy following surgical amputation for the primary lesion. The chemotherapy regimen utilized vincristine, actinomycin D, and cyclophosphamide (in high pulse doses) administered intermittently over a 55-week period. Four of the 12 patients (33%) are surviving with no evidence of disease 54 months or longer from time of surgery. These results suggest that the 3-drug combination used in this study should be considered in the formulation of future multidrug adjuvant chemotherapy programs in the management of patients with osteosarcoma.


Implications of genetic variation in mouse brain structure for electrode placement by stereotaxic surgery. The spatial locations of several forebrain and midbrain fibre tracts have been compared across seven commonly used inbred, hybrid and outbred mouse strains using a series of electrolytic lesions of known positions with respect to a stereotaxic instrument. Highly significant genetic variation was found in the position of lambda, bregma and several major fibre tracts with respect to interaural zero; in the locations of fibre tracts with respect to bregma; and in the location of fibre tracts with respect to each other. It was demonstrated that stereotaxic coordinates of a structure for one strain could not be used for other strains. Errors resulting from improper alignment of the head in the instrument and histological artifact were also determined. These errors showed no significant group differences, but they revealed that within-group variation was substantially affected by such imperfections.


Penfluridol: a long-acting oral neuroleptic. A controlled study. The clinical promise of penfluridol as a long-acting oral antipsychotic medication has led to a number of controlled studies designed to verify its usefulness. These studies have been reviewed and compared with a controlled study carried out by the authors. The data obtained from this study have tended to confirm the impression of previous investigators that penfluridol, administered in a dosage of 40 to 80 mg on a once-a-week basis in a single dose, compares favorably with the antipsychotic activity of those neuroleptics requiring administration on a daily basis.


Crossover clinical trial of benapryzine and trihexyphenidyl in Parkinsonian patients. A crossover clinical trial of benapryzine and trihexyphenidyl in ten parkinsonian patients during a four-month study is described. The improvement from initial disability ranged from 29.5 to 64.4 per cent for benapryzine and from 27.2 to 64.9 per cent for trihexyphenidyl. The effect of benapryzine and trihexyphenidyl on most parkinsonian symptoms did not differ significantly. Patients taking benapryzine had significantly fewer of the common side effects of trihexyphenidyl but more sialorrhea. It is uncertain whether benapryzine is superior to trihexylphenidyl in the treatment of Parkinson's disease.


Guidelines for diet counseling.

Guidelines for diet counseling. Members of the American Dietetic Association have shown an interest in having some procedural guidance in the process of dietary counseling. Such guidelines, prepared by a committee of the Diet Therapy Section, are presented. They include: a clarification of terminology; requirements for counseling in terms of knowledge, skills, and equipment; recommended procedural steps in counseling; and a supportive bibliography. While these guidelines are intended chiefly for dietitians in clinic and/or ambulatory care situations, the principles, philosophy, and general methodology should be basic to any educational situation, recognizing that circumstances can well present varying limitations.


Effects of protriptyline and perphenazine in neurotic depressed outpatients. A four-week comparison of protriptyline 10 mg three times a day and perphenazine 2 mg three times a day, alone and in combination, and a placebo in a group of nonpsychotic depressed outpatients showed no significant differences among any of the treatment groups after both two and four weeks, and a slight trend in favor of the placebo group. A significant positive correlation was observed between changes in hostility and changes in depression, contrary to many commonly held assumptions regarding the relationship between hostility and depression.


The pregnant adolescent. A review of nutritional research and programs. Avialable nutritional research on pregnant adolescents shows that suboptimal food intake is common. Multiple social, psychologic, and economic problems may mitigate against attainment of adequate nutritional status. Use of supplements will raise the level of some nutrients, but in reported studies, their use by girls was erratic, and routine use is not favored by many practitioners. Results of nutritional counseling are mixed. Multidisciplinary programs, usually providing comprehensive prenatal care while enabling girls to continue their education, have been instituted in many areas, reflecting a changed attiude of school officials toward the pregnant adolescent. Most of these programs include the nutrition classes, counseling, or services valuable to mother and child; however, they do not reach all pregnant adolescents and are not a panacea guaranteeing adequate nutrition for them. More research and a national policy to provide optimal nutrition for all pregnant women, including the adolescent, are needed.


Ibuprofen versus buffered phenylbutazone in the treatment of osteoarthritis: double-blind trial. In a double-blind multiclinic trial, a new nonsteroidal anti-inflammatory agent (ibuprofen) was compared with an established therapeutic agent (phenylbutazone-alka) for the treatment of osteoarthritis. Of the 159 patients from the 17 contributing clinics, 144 completed the four weeks of therapy. More than 60 per cent of them reported improvement in exercise-related pain by week 4, and there was no significant difference between treatment groups. The patients' and the physicians' evaluations of the total state of disease, as well as range-of-motion and functional tests, demonstrated similar degrees of improvement in both treatment groups. The incidence of side effects was within acceptable limits, and the frequency distribution was similar in both groups. Of the 70 reported side effects, 29 were considered by the investigator (blind trial) to be drug-related-11 in association with ibuprofen and 18 with phenylbutazone-alka. Hematologic and blood chemical studies, as well as urine and stool examinations, yielded normal results with the exception of a reduced mean value for serum uric acid and a slightly elevated mean value for SGPT in the phenylbutazone-alka group.


Effects of a procaine preparation (Gerovital H3) in hospitalized geriatric patients: a double-blind study. The effects of Gerovital H3 (a specially stabilized form of procaine hydrochloride) on geriatric psychiatric patients were assessed in a double-blind study at Bronx State Hospital. The mean age of the subjects was 73 years and the average rating for the severity of organic symptoms was "moderate." During the first six weeks of study, the patients were each given a 5-ml injection of either Gerovital or placebo (saline) intramuscularly three times a week. This dosage was doubled to 10 ml per injection during the second six weeks. Nine Gerovital and 10 control subjects completed the first six weeks; and 6 Gerovital and 7 control subjects completed the entire 12-week study. Objective rating scales were used to evaluate patients on measures of interpersonal functioning, cognitive ability, psychiatric symptoms, and urine and blood chemical findings. All subjects were assessed before treatment and at six weeks and twelve weeks of the study. Side effects were recorded at two-week intervals. On most measures the variability between subjects was quite large, whereas differences between average scores for the two groups usually were small The few significant differences showed no systematic pattern and would be expected to occur by chance alone when so many statistical comparisons are made. The overall results of this double-blind study strongly indicated that, among these hospitalized geriatric patients with organic symptoms, Gerovital H3 had no ameliorative effect on either psychologic or physiologic functioning.


Treatment of trigeminal neuralgia in the aged by a simplified surgical approach (percutaneous electrocoagulation). Trigeminal neuralgia can be excruciatingly painful. For aged patients, among whom trigeminal neuralgia most commonly occurs, medical therapy may often afford relief of pain. However, since this disorder is likely to become worse with increasing age, medical therapy eventually may fail because of increasing drug requirements and intolerable side effects. Major forms of surgical treatment requiring craniotomy may afford longstanding relief but are associated with significant side effects in 3-5 per cent of cases. For these reasons the authors began to test a new surgical approach, i.e., stereotaxic percutaneous electrocoagulation of the trigeminal nerve. More than 300 patients have been treated by this technique in the past five years. Successive radiofrequency lesions allow the production of a graded sensory deficit sufficient to relieve pain while preserving touch and motor function in the face. Unnecessary suffering from the severe pain of trigeminal neuralgia need not occur because of age or a debilitated condition which might ordinarily preclude surgical treatment. More than 100 patients over 65 years of age (4 of them older than 90) have been treated. Of this group, 93 per cent reported good to excellent results. Seven per cent have had some recurrent pain during the follow-up period; however, recoagulation has been required in only 5 per cent of these patients. Recoagulation may be readily performed in any patient who shows evidence of fading of the sensory deficit. The electrocoagulation technique and the results in elderly patients are discussed.


The water agar test: a new test to measure the bacteriological quality of cream. A new test, the water agar test, is described that gives a qualitative index of the presence of bacteria that indicate contamination of the cream, poor storage conditions or both of these factors. The method is simple and requires little equipment. The bacteria grow in a film of diluted cream adsorbed on the surface of a non-nutrient base. After incubation at 30 +/- 0-5 degrees C. for 18-20 hr., a proteolytic and mucoid colony count is obtained which has the same percentage coefficient of variance as a standard plate count. An examination of the effect of storage at different temperatures on the types of bacteria present in cream showed that of all the tests done initially, only the water agar test could predict subsequent bacterial growth with any consistency. The multiplication of presumptive coliform organisms occurred even at 3-5 degrees C. Irrespective of the colony count, the methylene blue reduction time was not shorter than 7 1/2 hr. unless the bacteria were in the logarithmic phase of growth when sampled. A survey was made of the bacterial flora of 188 retail samples of double cream of 15 different brands. The age of the samples varied from freshly separated cream to cream that had been kept in the shop for a day longer than that recommended for sale. The water agar test was compared with the colony count, the presumptive coliform test, a confirmatory coliform count in violet red-bile agar, a lipolytic colony count, a staphylococcal count and the methylene blue reduction test.


Quality control tests of two salmonella enrichment media using different inocula. Inocula of polluted water naturally infected with salmonellas effectively distinguished six brands of selenite and six brands of tetrathionate enrichment media into satisfactory and unsatisfactory categories. Minimal inocula of pure cultures differentiated the tetrathionates, but not the selenites. Inocula of naturally infected chicken giblets suggested that there was a difference between two comparable brands of tetrathionate, but this was not statistically significant. The difference was, however, clearly demonstrated by minimal inocula of pure cultures. Intensive investigation of two inferior tetrathionates revealed inhomogeneity in the distribution of brilliant green in one bottle of one brand. The importance of the salmonella serotype and even the colonial variant used for the pure culture inoculum was also demonstrated.


Bacteriological tests as indices for the development of off-flavours in cream. Bacteriological tests alone are not capable of predicting the keeping quality of cream, since taints can develop which are due to non-bacterial action. When spoilage is due to bacterial growth, the water agar test was found to be more accurate than other tests that were examined.


Citrobacter koseri. I. An extended antigenic scheme for Citrobacter koser (syn. C. diversus, Levinea malonatica). The names Citrobacter koseri, Levinea malonatica and Citrobacter diversus are synonymous. As a result of the study of representative strains of these organisms an antigenic scheme comprising seven O antigens has been proposed. The examination of a further 165 strains from clinical sources suggests that an additional seven antigens should be included in the scheme. The serological and biochemical reactions of the test strains for all 14 O antigens are described.


The effect of thermal pollution on the distribution of Naegleria fowleri. The distribution in the environment of Naegleria fowleri, the causal agent of primary amoebic meningoencephalitis has been investigated in this study. N. fowleri was isolated only from a thermally polluted canal. These amoebaflagellates were not isolated from another thermally polluted canal in the neighbourhood indicating that, apart from high temperature, other factors are involved in the selective proliferation of N. fowleri. This species was absent in all other samples originating from two canals, a stream, two lakes, several reservoirs and slow sandfilters of a water supply service and also a water distribution network. Many other amoebae able to grow at 42 degrees C. were found in different places. Most of the N. fowleri strains isolated were not virulent for mice, although they showed all the characteristics of the pathogenic strains.


Prolongation of allograft survival in mice by administration of anti-Thy 1 serum. I. Mediation by in vivo activation of regulatory T cells. A single 0.5-ml dose of anti-Thy 1.2 serum raised in AKR (H-2k, Thy 1.1) mice by immunization with C3H (H-2k, Thy 1.2) thymocytes prolonged mean graft survival (MGS) of C57BL/L (H-2b, Thy 1.2) skin on BALB/c (H-2d, Thy, 1.2) mice from 9.2 to 13.0 days (p less than 0.01). Absorption of anti-Thy 1.2 serum with C3H but not AKR brain removed this activity. MGS was prolonged by anti-Thy 1.2 serum given as long as 2 weeks before grafting. Four separately raised anti-Thy 1.2 preparations prolonged MGS, one to a dilution of 1:128. Anti-Thy 1.1 serum prolonged MGS on AKR mice but not on BALB/c mice. Thymectomy 3 weeks or more before graft placement or treatment of BALB/c mice with anti-lymphocyte serum (ALS) diminished the effectiveness of anti-Thy 1.2 serum. Furthermore, infusion of 25 times 10(6) spleen cells from BALB/c mice given anti-Thy 1.2 serum prolonged MGS on syngeneic recipients to 15.6 days whereas spleen cells from mice given no treatment or ALS had no effect. Thymocytes from anti-Thy 1.2 serum treated donors were ineffective in prolonging MGS on transfer. Transfer or serum from anti-Thy 1.2 serum-treated mice also had no effect. The transfer of graft prolongation with lymphoid cells from anti-Thy 1 serum treated mice suggests anti-Thy 1 serum may prolong MGS by activating suppressor T cells.


Endotoxin-stimulated spleen cells: characterization of the responding cells. The percentage of endotoxin-stimulated lymphoblasts possessing C3 receptors (CR) decreases during a 72-hr period of in vitro cultivation. In studies reported here we attempted to determine whether the drop in the percentage of CR+ blast cells resulted from receptor loss by transformed cells or whether it resulted from the proliferation of additional blast cell types which lacked the CR. Using methods which permitted simultaneous detection of cytoplasmic IgM, the presence of the CR, and 3H-thymidine uptake we found that: a) LPS stimulation resulted in a progressive increase in the number of IgM+ blasts, b) these constituted approximately 30% of the total cell population at 72 hr after LPS stimulation, c) of the IgM+ blasts 72 to 84% were CR+ and 16 to 28% were CR- at 72 hr, d) from 48 to 72 hr there was no decrease in the percentage of IgM+ blasts possessing the CR, and e) an additional blast cell type was observed which incorporated 3H-thymidine but which was IgM- and lacked the CR. These cells constituted from 11 to 27% of total 3H-thymidine+ cells at 72 hr after LPS stimulation. We conclude that the decrease in the percentage of CR+ blasts in LPS-stimulated spleen cell cultures is not due to receptor loss from IgM+ blasts cells but apparently is partly due to the stimulation and proliferation of other cell type(s) which lack(s) the CR. This cell(s), although its functional identity is unknown, does not appear to be of thymic origin. The heterogeneity of the markers on LPS-stimulated blast cells suggests that a heterogeneous population of cells responds to LPS.


Studies on leukocyte migration inhibiton: the role of E rosette-forming cells in specific antigen-induced inhibition of migration. The role of E rosette-forming cells in leukocyte migration inhibition was investigated with human buffy coat cells fractionated by combined E rosette formation and Ficoll-Hypaque (FH) density gradient centrifugation. The migration of unfractionated dextran-sedimented leukocytes and FH-separated mononuclear cells (MN) were equally inhibited in the presence of Escherichia coli somatic antigen (E. coli), purified protein derivative of tuberculin (PPD) and influenza virus antigen (Flu). Polymorphonuclear leukocytes (PMN) separated from whole leukocytes by one FH density gradient fractionation contained greater than 1.0% E rosette-forming cells and were inhibited by E. coli and PPD. PMN submitted to three transits of FH (DF III) contained less than or equal to 0.02% E rosette forming cells and were unaffected by each of the three antigens. The migration of mixtures of 10% MN and 90DF III cells were inhibited in the presence of specific antigen, while MN depleted of E rosette-forming cells (less than or equal to 0.5%) pooled with equal numbers of DF III cells did not respond to any antigen by migration inhibition. These results show that PMN purified by three transits of FH are unable to respond to specific antigen by inhibition of migration from microcapillary tubes, and specific antigen-induced inhibition of migration of peripheral blood leukocytes does not occur in the absence of D rosette-forming cells in normal subjects.


Kinetic study of T lymphocytes after sensitization against soluble antigen. I. Separation on density gradients. Sensitized T cells in inguinal lymph nodes (LN) of rats immunized with ovalbumin and adjuvant can be separated on BSA density gradients from T cells responding to Con A and PHA. Further, these OA-sensitized T cells are separable from B memory cells responsive to OA. On days 9 and 17 after sensitization, OA-reactive cells peak and are found in enriched numbers in the lighter bands A + B (B-band), and C, respectively whereas Con A and PHA reactive cells, on both days, are concentrated in bands C and D, respectively. On day 9 at least 20 to 30% of B-band cells can reinitiate DNA synthesis and produce high concentrations of lymphotoxin in response to OA in culture, but show only a marginal blastogenic response to Con A or PHA and cannot produce lymphotoxin in response to PHA. These cells are defined as OA-sensitized T cells because a) treatment with rat isoantiserum specific for T lymphocytes plus complement abolished their ability to incorporate 3H-thymidine in response to OA and Con A or PHA; b) membrane-bound immunoglobulin could not detected with indirect immunofluorescence staining; c) histologic studies demonstrate massive hypertrophy of thymus-dependent paracortical areas and absence of germinal centers in inguinal LN at this time, and d) they produce lymphotoxin after challenge with OA, but not with PHA, whereas cells of the other bands produce lymphotoxin in response to PHA, but not to OA. Addition of purified macrophages to the various cultures did not affect the levels of response observed. Day 17 C-band cells are regarded as B-memory cells because treatment with T-specific antiserum plus complement enhanced their response to OA, many germinal centers now are present in inguinal LN, and a significant proportion of cells of bands B and C, negative on day 9, show positive immunofluorescence staining at this time.


Physicochemical characterization of chemotatic lymphokines produced by human T and B lymphocytes. Ostensibly pure populations of T and B lymphocytes were separated from peripheral human blood by a sequential rosetting method. These cells, and mononuclear leukocyte (MNL) cell preparations (60 to 85% T cells), were cultured in vitro and the supernatants of these cultures were assayed for chemotactic lymphokine (CTX) activity. B lymphocytes were activated to produce CTX by binding erythrocytes sensitized with antibody and complement (EAC) to their C3 receptors and phytohemagglutinin (PHA) stimulation of isolated T cells was used to produce T cell-derived CTX. Sephadex G-100 gel filtration chromatography of supernatants obtained from T and B cell cultures showed that both cell types produce a CTX of approximate molecular weight 12,500 daltons. This result is identical to that previously obtained in characterizing the CTX produced by buffy coat leukocytes. Isoelectric focusing of lymphocyte supernatants and chemotactically active Sephadex G-100 fractions showed that the CTX produced by B cells and PHA-stimulated MNL contained two peaks of lymphokine activity, a major peak at pH 10.1 and a minor peak at pH 5.6. Additionally, these studies show that the chemotactic complement peptide C5a has an isoelectric point of pH 8.7 and is antigenically distinct from T and B cell-derived CTX, although relatively homogenous by gel filtration, contain two isoelectrically distinct species. In addition, these findings suggest that T and B cells although possessing different membrane receptors and requiring different mechanisms of activation to initiate lymphokine synthesis, produce lymphokines with identical physiocochemical properties.


Isolation and characterization of the sheep erythrocyte receptor in man. Properties of the sheep erythrocyte receptor on human lymphocytes were examined from two different aspects: inhibition of E-rosette formation by antisera, and extraction of T and non-T cells with 3 M KCl. Antisera to several T cell populations produced significant inhibition of E-rosette formation by human peripheral blood lymphocytes, whereas anti-B cell, anti-beta2-microglobulin, anti-I, and anit-i were ineffective. Sera with activity to HL-A subgroups on the HPBL tested produced complete rosette inhibition, although nonspecific anti-HL-A produced no inhibition. T cell preparations extracted by a 3M KCl technique contained activity that could be bound to SRBC and produce inhibition of rosette formation by HPBL. This inhibitory component could be quickly eluted from the SRBC and retain its inhibitory activity. No inhibitory activity could be demonstrated in extracts of B cells and non-lymphoid cells. These results suggest that 1) T cell-specific and HL-A specific antisera inhibit E-rosette formation; 2) beta2-M is not involved in the SRBC receptor as an active component; and 3) the receptor may be isolated by 3 M KCl extraction, and partially purified by the use of SRBC as a specific immunoadsorbent, without loss of biologic activity.


Lymphocyte abnormalities in Aleutian disease virus infection of mink: decreased T lymphocyte responses and increased B lymphocyte levels in persistent viral infection. Persistence of virus and a marked hypergammaglobulinemia are consistent features observed in mink with Aleutian disease virus infection. These events may result from altered lymphocyte function in infected mink. To test this possibility we determined the ability of lymphocytes from infected mink to respond to phytolectins (PHA and Con A), and a specific antigen (KLH) in a lymphocyte transformation test and quantitated the number of lymphocytes bearing surface immunoglobulin. Infected mink had a significantly lower response than normal mink to PHA and Con A (P less than 0.025), and the difference in the responses increased progressively with time post-infection. Infected mink immunized with KLH also had a decreased ability to respond to this antigen when compared to normal immunized mink (P less than 0.001). The number of lymphocytes bearing surface immunoglobulin was significantly elevated in three of 10 mink infected for 10 weeks, and in all of seven mink infected 32 weeks (P less than 0.001).


The expression on mouse lymphoid cells of Th-B, an antigen common to mouse B cells and thymus cells. A new cell-surface antigen of mouse lymphocytes, which was found earlier on normal and malignant plasma cells and about half the normal thymus cell population, has been shown to be on mature B cells and on the least immunocompetent subpopulation of cells of the thymus. The antigen was first detected by the cytotoxic effect on relevant cells of an in vivo purified rabbit antiserum raised against cells of the mouse IgM-producing plasma cell tumor MOPC-104E. We have now used a cell-transfer method to examine the effect of this antiserum (RantiM104E) on immunocompetent lymphocytes of spleen, including separated splenic B and T cells and thymus and bone marrow cells. We have found the antigen on the immunocompetent B lymphocytes of the spleen, but not on any other of the immunocompetent cells examined. The method involves determining the effect of the antiserum on the development of plaque-forming cells in the spleens of irradiated mice that have received lymphocytes from immune or non-immune donors by treating the lymphocyyes  with RantiM104E antiserum before transfer. Plaque-forming cells are suppressed when antiserum-treated spleen cells are transferred to the irradiated mice.This suppression was found to be due to action on only the splenic B cells. The helper activity of splenic T cells was not reduced significantly by antiserum treatment. Neither was the helper activity of thymus cells similarly treated reduced by the antiserum. We found, in fact, that the subpopulation of the thymus cells which is resistant to the antiserum was more immunocompetent than the total thymus cell population. In addition, the hydrocortisone-resistant thymus cells were also found to be resistant to the cytotoxic action of the antiserum. The antiserum had no effect on bone marrow cells in the cell-transfer procedure. The antigen involved, which we are designating "Th-B" appears to be a B cell line marker which appears relatively early during the differentiation of mouse B cells from precursor cells and is lost during maturation of T cells.


Antigenic competition between heterologous erythrocytes in mice: interference with competition-inducing effect of sheep erythrocytes by inclusion of non-competing antigen. Antigenic competition was studied on reciprocal combinations of non-cross-reacting erythrocyte (RBC) antigens, i.e., sheep erythrocytes (SRBC), horse erythrocytes (HRBC), and chicken erythrocytes (CRBC), as competing antigen or as test antigen. Unlike SRBC and HRBC, CRBC did not induce antigenic competition, and mice previously injected with CRBC showed normal direct and indirect hemolysin plaque-forming cell (PFC) responses to the other RBC subsequently injected. Formaldehyde-treated (F) SRBC that retained the capacity to prime mouse for secondary anti-SRBC antibody response but lost the capacity to elicit primary antibody response also could not induce antigenic competition. CRBC or F-SRBC, i.e., non-competing antigen, included with SRBC interfered with the competition-inducing effect of the latter RBC. When mice that had been injected with SRBC together with either one of the non-competing antigens were subsequently immunized with HRBC their anti-HRBC PFC responses were significantly restored, whereas mice pre-injected with SRBC alone showed markedly suppressed PFC responses to HRBC. Although the non-competing antigen was found not to affect both the direct and indirect PFC responses to the simultaneously injected SRBC, the response of the immune system to the non-competing antigen should alter the concurrent response to SRBC, and probably should prevent the generation of the suppressor thymus-derived (T) lymphocytes, allowing the generation of the helper T lymphocytes for IgM and IgG anti-SRBC antibody formation.


Antigen-initiated B lymphocyte differentiation. IV. The adherence properties of antibody-forming cell progenitors from primed and unprimed mice. Distinct sub-populations of B lymphocytes may be fractionated by passage through glass-bead columns, due to a temperature-independent adherence of certain cells. The relative adherence properties of AFC-progenitors from spleens of primed or unprimed CBA mice were compared with an adoptive immune assay employing as antigen the hapten NIP on the carrier polymerized flagellin, under conditions where T lymphocytes did not limit the response. The AFC-progenitors for an anti-NIP IgM response from unprimed mice, whether conventional, germ-free, or athymic were more adherent than the majority of cells in the spleen. The AFC-progenitors for an IgG response from hapten-primed mice were less adherent that the majority of spleen cells. The results indicate that there are sub-classes of AFC-progenitors, differing in surface properties and probably corresponding to differences between the "antigen-inexperienced" and the "memory" B cell populations.


Antagonism of B lymphocyte mitogenesis by anti-immunoglobulin antibody. Anti-mouse-immunoglobulin-antibodies (AMIg) were shown to suppress mitogenesis in B lymphocytes that were cultured with lipopolysaccharide, (LPS), or polymerized-flagellin, (POL). Preincubation of spleen cells with AMIg plus LPS has a greater suppressive effect on a subsequent mitogenic response to LPS than did preincubation with AMIg alone. Preincubation with LPS alone had, if anything, a slight enhancing effect. These experiments suggested that AMIg did not suppress mitogenesis by interference with the initial binding of the mitogen, but rather exerted a separated negative effect that occurred subsequent to, or in the process of, B cell-stimulation.


IgG antibody suppression of the IgM antibody response to Toxoplasma gondii in newborn rabbits. Studies were performed in an experimental rabbit model in an attempt to explore the possibility that the absence of an IgM antibody response in human infants congenitally infected with Toxoplasma may be caused by a suppressive effect of IgG antibody passively transferred from the mother. The IgM antibody response to Toxoplasma gondii infection in newborn rabbits previously injected with IgG Toxoplasma antibody was significantly suppressed as determined by an immunofluorescent antibody assay (IgM-IFA). In newborn rabbits infected with Toxoplasma, IgM-IFA antibody was first demonstrated 15 days after infection and reached a peak titer of 1:128 in two animals on days 20 and 25. In newborn rabbits injected with IgG Toxoplasma antibody and then infected with Toxoplasma, the Igm-IFA antibody and then infected with Toxoplasma, the Igm-IFA antibody response was considerably lower; the highest titer was 1:16 and occurred 25 and 30 days after infection. The results of these studies lend support to the concept that maternally transmitted IgG may suppress the IgM antibody response in the fetus and newborn infant.


Pretreatment of murine thymocytes by phytohemagglutinin inhibits the binding of H-concanavalin A. The binding of H-concanavalin A (Con A) to cortisone-resistant and normal mouse thymocytes, which greatly differ in mitogenic responsiveness, was compared. Almost equal numbers of 3H-Con A molecules (7 x 10(5) molecules per cell) were found to bind to both cell types, and the rates of H-Con A binding were nearly identical. The binding capacity of thymocytes for H-Con A decreased rapidly after removal from the thymus and incubation in protein free synthetic medium. Similarly, mitogenic responsiveness droped rapidly after cells were exposed to this medium. H-Con A molecules bound to cells were dissociated from them after a brief incubation, about 30 percent of radioactivity being lost in 2 hr with increasing radioactivity being found in the supernatant. When cells were pretreated with PHA-P, which is a T cell mitogen, their mitogenic responsiveness to Con A was suppressed along with H-Con A binding to the cells. In contrast, a B cell mitrogen, LPS, that has a synergistic effect on the Con A mitogenic response in thymocytes had no effect on binding of H-Con A to thymocytes.


Hyperacute rejection and enhancement of mouse skin grafts by antibodies with a distinct specificity. Pure 7S IgG2 was prepared from B6AF1 anti-B10.D2 ascites fluid by affinity chromatography. This preparation was extensively absorbed with B10.D2 red blood cells to remove the antibodies directed against the serologically defined antigens of the major histocompatibility complex. After absorption the serum had retained in vitro cytotoxic activity against 45 percent of B10.D2 spleen cells. The absorbed and unabsorbed 7S IgG2 antibodies were tested for their capacity to induce enhancement and, in the presence of rabbit complement, hyperacute rejection of B10.D2 skin grafts in B6AF1 recipient mice. The enhancing capacity of 7S IgG2 was completely unimpaired after absorption. However, the absorbed preparation was unable to induce hyperacute rejection of the grafts even if very high doses were administered. The results show that hyperacute rejection of skin grafts in this model is mediated by antibodies directed against the serologically defined antigens (i.e., H-2K antigens) of the H-2 complex. They further support the hypothesis that immunologic enhancement is mediated by antibodies directed against Ir-region-as-sociated antigens.


Cytotoxic activity of lymphocytes. V. Role of soluble toxin in macrophage-inhibited cultures of tumor cells. Macrophage cell-mediated cytolysis (M-CMC) of tumor cells may be measured by culturing target cells on macrophage monolayers and determining target cell survival by either Rb or H-thymidine incorporation. Using these techniques, one can demonstrate cytotoxicity of nonimmune human and rat macrophages to a variety of tumor cell lines. Toxicity becomes maximal 8 to 16 hr after attaching macrophages to plastic surfaces and disappears after 36 to 48 hr. Addition of Escherichia coli endotoxin at 48 hr restores macrophage toxicity. Culture of macrophages on plastic surfaces also results in the release, nonspecifically, of a soluble cytotoxin whose kinetics of release, disappearance, and reappearance after endotoxin all parallel the M-CMC. The amount of toxin released into cultures containing target cells is markedly reduced, suggesting that it is the mediator of the M-CMC and that it is utilized or absorbed during target destruction. Sephadex G-100 chromatography and treatment with heterologous anti-lymphotoxin can distinguish this toxin from lymphotoxin.


Genetic control of susceptibility to experimental allergic encephalomyelitis and the Ag-B locus of rats. Injection of guinea pig spinal cord with complete Freund's adjuvant induces EAE in Lewis (Ag-B1) and DA (Ag-B4) rats, but not in rats of the BN (Ag-B3) strain. When BN is crossed with Lewis, susceptibility is determined by a gene linked to the Ag-B histocompatibility locus, but a much more complex mode of inheritance is involved when BN and DA rats are crossed. Six inbred strains possessing the Ag-B1 histocompatibility allele have been found to be susceptible to EAE. Evidence for recombination between the Ag-B locus and the gene determining susceptibility to EAE has not yet been obtained.


Class differences in the antibody response of HLC and LLC mice to sheep red blood cells. The immune response to sheep red blood cells (SRBC) has been studied in two mouse lines selectively bred for a difference in leukocyte levels. Low leukocyte count (LLC) mice responded to multiple injections of 4 x 10(7) SRBC with a predominant IgM response whereas high leukocyte count (HLC) mice produced IgG almost exclusively, even after a single injection of SRBC. F1 mice responded like the LLC parent and backcross data implied genetic control at multiple loci. No correlation was found between the class of antibody produced in response to SRBC and the leukocyte count in backcross mice. Large numbers of plaque-forming cells (PFC) were detected in the spleens of HLC mice relative to the number detected in LLC spleens after primary and secondary immunizations with SRBC.


T cell activation and cellular cooperation in autoimmune NZB/NZW F hybrid mice. Old (6 months) overtly autoimmune female NZB X NZW F1 (B/W) mice were markedly hyporesponsive to sheep erythrocytes (SRBC). The response to SRBC was restored by a) simultaneously injecting lipopolysaccharide (LPS) with SRBC or b) transferring bone marrow and thymocytes with SRBC into lethally x-irradiated (100 R) syngeneic old recipients. The in vitro PFC response of old B/W spleen cells to SRBC was restored by adding in culture a) theta-positive and radioresistant spleen cells from old B/W mice primed with homologous antigen or b) activated T cells from the spleens of lethally x-irradiated (1000 R) old B/W mice injected with old syngeneic thymocytes and SRBC but not horse erythrocytes. Various populations of unprimed lymphoid cells from young (4 to 6 weeks) female B/W mice, which respond normally to SRBC, were not capable of restoring the response of old syngeneic mice in vitro or in vivo. These results suggest the existence of a suppressor of T cell activation and/or B and T cell interaction in old autoimmune B/W mice.


Inhibition of in vitro lymphokine synthesis by glucocorticosteroids. At pharmacological levels, glucocorticosteroids inhibited two antigen-induced lymphocyte functions, in vitro proliferation and lymphokine synthesis. Lymphocyte production of both macrophage chemotactic factor (CTX) and macrophage inhibition factor (MIF) were decreased in the presence of hydrocortisone. The corticosteroid also blocked the action of MIF on macrophages but did not interfere with the action of CTX on macrophages. Thus, steroids can suppress the immune response at two different stages: by blocking lymphocyte activation and mediator synthesis and also by interfering with the interaction of certain effector molecules with their target cells. Furthermore, these findings suggested that MIF and CTX have distinct mechanisms of action on the effector macrophages, one being sensitive and one resistant to steroids.


Antigenic selection of IgG precursor subpopulations. The temporal increase in the antigen-binding characteristic of the IgG antibody synthesized after primary immunization is thought to result from antigen preferentially selecting those precursors of the antibody-forming cells with higher affinity receptors. We have investigated this regulatory role of antigen on the selection and expression of IgG precursor subpopulations in rabbit lymph node populations undergoing the secondary anti-bovine serum albumin (BSA) response in vitro. The data indicate that although immunologic memory increased between 1 and 4 months after immunization there was not an increase in antigen sensitivity. We also determined the avidity of the IgG antibody secreted by PFC elicited in the cell cultures. Although the avidity of antibody secreted by PFC elicited with low concentrations of antigen was 2-fold greater than when high concentrations of antigen were employed, analyses of the avidity subpopulations of IgG PFC revealed that memory potential was not restricted to high affinity-primed cells. Moreover, populations of PFC that gave differing avidity values were similar in terms of their plaque diameters and IgG secretion rates.


The enhancing effect of mitogens on the in vivo immune response of rabbits. The effect of Con A and PHA on the primary humoral immune response in rabbits was investigated and compared with the effect of LPS and irradiation. Both Con A and PHA enhanced PFC levels and precipitating antibody titers to HuIgG when injected simultaneously with AHuIgG. Comparable enhancement of the response to HuIgG was obtained by simultaneous injection of LPS with AHuIgG or by irradiating rabbits (500 R) 24 hr after injection of the antigen. Further studies with Con A showed that its adjuvant effect was not restricted to a particular antigen since it also enhanced antibody responses to both SRBC and Bov Tg when injected simultaneously with either of these antigens. Although optimal enhancement of the response to HuIgG occurred when antigen and mitogen were injected simultaneously, some enhancement was obtained if Con A was injected 2 days after antigen, whereas the response was decreased if Con A was injected 2 days before antigen. Splenic PFC in A/J mice injected with doses of AHuIgG and Con A equivalent on a body weight basis to those used in rabbits were also enhanced by simultaneous injection of Con A with the antigen.


The effect of staphylococcal enterotoxins on the primary in vitro immune response. Staphylococcal enterotoxins, like several plant lectins, have been shown to stimulate mitogenic activity and lymphokine production in lymphocytes. The effect of enterotoxins A and B on the primary in vitro plaque-forming cell (PFC) response of mouse (C57BL/6) spleen cells to sheep red blood cells (SRBC) was examined. PFC responses in 4- or 5-day cultures were inhibited greater than 90% by 0.1 mug enterotoxin A and by 3- to 6-mug enterotoxin B when the toxins were added to the cultures either at the time of SRBC addition or 24 hr before. Both enterotoxins A and B were shown to be T lymphocyte mitogens. Kinetic data indicated that the enterotoxins (10 mug) inhibited the PFC response as early as day 3, but were relatively more inhibitory at days 4 and 5. The PFC response was inhibited when enterotoxins were added to cultures at the same time as antigen or a day later; it was enhanced when added at day 2; and it was unaffected when added at day 3 of 4-day cultures. Further, the presence of enterotoxin A during the first 24 hr of culture and subsequent removal was still as effective in inhibiting the PFC response as when it was present throughout the culture period. The PFC inhibitory properties of enterotoxins appear, then, to affect some early event(s) in the in vitro immune response. Alpha-methyl-D-mannopyranoside blocked the PFC inhibitory effect of concanavalin A (con A), but it had no effect on enterotoxin A. The two mitogens appear, then, to react with different receptors on the lymphocytes or to differ in the dynamics of their reactivity. The effects of staphylococcal enterotoxins on the PFC response of spleen cells were remarkably similar to those reported for lectins such as con A. Enterotoxins are structurally simpler than con A and should therefore be quite useful in studying various biologic activities of lymphocytes.


Ia alloantigen(s) detected on thymocytes by use of a fluorescence-activated cell sorter. With the increased sensitivity of detection of fluorescence made possible by a fluorescence-activated cell sorter (FACS), Ia antigen(s) have been demonstrated, although very small amounts, on murine thymocytes by using an indirect immunofluorescent technique. Absorption studies indicate that the Ia antigens detected on thymocytes are also present on spleen B cells, however, they do not rule out the possibility that certain Ia antigens may exist only on B cells. These studies suggest that the surface distribution and/or the decreased density of Ia antigens on thymocytes relative to peripheral B cells may preclude their demonstration by more conventional assays. Although these studies demonstrate Ia antigens on less than 50% of thymocytes, the percentage may in fact be much higher since current methods do not allow an upper limit to be assigned.


Antigenic competition between two sequentially acting antigens. Immunosuppressive effect of T cells in spleen and lymph nodes of mouse. This study concerns the antigenic competition among heterologous red cells, a T suppressor-dependent phenomenon, in three cases, when the two antigens are administered by various routes, in (a) splenectomized mice or in Nude mice reconstituted with (b) lymph node cells, or spleen cells. Competition occurs whatever the routes of antigenic introduction, except when the two antigens are injected subcutaneously in the same leg and the anti-SRC plaque-forming cells are counted in the draining lymph nodes. In that case, the first antigen plays the role of adjuvant to the second. An adjuvant effect is also observed in splenectomized mice when the antigens are introduced intraperitoneally. However, competition does not occur in Nude mice but is present when those animals have been reconstituted either with lymphnode cells or with spleen cells. Those results play in favor of the presence of T suppressors, both in lymph nodes and in spleen; however, in lymph nodes, the immunosuppressive action induced by the first antigen is exceeded by an concomitant adjuvant effect.


Separation of lymphocytes by specific adherence to cellular monolayers containing protein A of Staphylococcus aureus. A technique was developed for the isolation of lymphocytes according to their surface antigenic markers. The method was based on the general reaction of protein A (SpA from S. aureus) with the Fc-part of an IgG antibody. Monolayers of S. aureus or Spa-coated sheep red blood cells (SpA-SRBC) fix antibody-charged cells specifically; non-fixed cells are easily removed by washing. Alternatively, the monolayers can be treated with a cell surface specific antibody prior to addition of non-charged cells. Monolayer-adhered cells were detached by the addition of lysostaphin (bacterial monolayer) or ammonium chloride (SpA-SRBC monolayer). As an example, Ig-bearing cells were isolated from mouse spleen lymphocytes using a specific rabbit anti-mouse Ig serum for charging either the cells or the monolayers. The recovery of Ig-bearing cells was approximately 84%. The purity of the cells was approximately 83% and the viability 89%. Antibody-SpA complexes on the surface of isolated cells were removed either by trypsin treatment or by cultivation. Ig on the cell surface is restored after 16 hr of cultivation.


B lymphocyte differentiation induced by lipopolysaccharide. I. Generation of cells synthesizing four major immunoglobulin classes. Cultured cells from mouse thoracic duct, spleen, lymph node, Peyer's patches, and bone marrow gave rise to cells containing large amounts of cytoplasmic IgM, IgGl, and IgG2 when stimulated by bacterial lipopolysaccharide (LPS). Differntiation of IgA producers occurred in bone marrow only, and to a lesser extent than other classes. Significant IgM responses preceded development of cells containing other classes. The differentiation of IgG and IgA producers did not appear to depend on T cells, since cultures from nu/nu or thymectomized-irradiated, bone marrow-protected mice responded as well as normals. Cultures from mice rendered deficient in B cells by anti-mu treatment responded normally to T cell mitogens, but did not proliferate or give rise to immunoglobulin-secreting cells when stimulated with LPS. Bone marrow cultures gave relatively meager proliferative responses to LPS, but generated as many or more immunoglobulin-secreting cells as did other tissues.


Sera and the in vitro induction of immune responses. I. Bacterial contamination and the generation of good fetal bovine sera. More than 200 samples of pooled fetal bovine sera (FBS) were tested to determine their capacity to support primary immune responses by cultured mouse spleen cells. Less than 10% of the FBS samples were fully supportive; the majority of the samples were moderately to very deficient. Several lines of evidence suggest that bacterial contamination during the processing of sera plays a major role in determining which sera are supportive. Serum samples known to have been temporarily contamined during processing were strongly supportive. Samples of FBS which were likely never to have been contaminated were deficient. Bacteria from a specific lot of supportive serum converted a very deficient serum into one which was supportive. Several mechanicsms by which bacteria could effect sera are discussed.


Sera and the in vitro induction of immune responses. II. Inhibitory effects of newborn calf sera on primary humoral responses. The effect of newborn calf (NBC) sera on the in vitro generation of immune responses were studied. NBC sera inhibited the generation of primary humoral responses to SRBC. The inhibitory activity was dose-dependent and diluted out rapidly. Adding inhibitory doses of NBC serum to test cultures at zero or 24 hr resulted in approximately the same amount of inhibition. Pulsing experiments showed that the test cultures were most sensitive to the inhibitory effects of NBC serum during the 24- to 48-hr period. In contrast to its inhibitory effects on primary humoral responses, NBC sera supported the in vitro generation of secondary humoral responses and significantly increased the responses of some of these over control levels. We found that sera obtained from newborn calves before nursing were not inhibitory for primary responses but sera obtained from the same calves 7 days later did inhibit these responses. We postulate that the inhibitory effects of NBC sera may represent a physiologic control mechanism which becomes active when immunologic maturity develops and that it possibly involves the inactivation of cell mediators.


Studies on the human T lymphocyte population. III. Synthesis and release of the lymphocyte receptor for sheep red blood cells by stimulated human T lymphoblasts. The IgG fraction of a goat antibody specific for human T lymphocytes (ATG) and that of a rabbit antibody directed (anti-Ts) were used to study the synthesis, display, and release of the receptor for SRBC (Ts) by normal human peripheral lymphocytes. A large percentage of lymphoblasts obtained by stimulation of lymphocytes with PHA or PWM formed multilayered rosettes with SRBC. The vast majority of these rosetting lymphoblasts were T lymphocyte derived, whereas few B lymphoblasts formed rosettes. Significant increases were found in the quantity of Ts on the lymphoblast membrane and in the supernatants from PHA-stimulated cultures, especially late in transformation. Furthermore, lymphocytes cultured with PHA and 3H-leucine synthesized and released radiolabeled Ts.


Studies on lymphokines: the production of antibody to guinea pig lymphotoxin and its use to distinguish lymphotoxin from migration inhibitory factor and mitogenic factor. Guinea pig lymphotoxin, which was produced by stimulating immune lymph node cells with the specific antigen, ovalbumin, was partially purified by the sequential use of Pevikon block electrophoresis, ion exchange chromatography on DEAE-cellulose, and polyacrylamide disc gel electrophoresis. The lymphotoxin recovered from the final step was purified approximately 200- to 300-fold as compared to the starting material, but nevertheless contained at least one major contaminant protein. Rabbits were immunized with the partially purified lymphotoxin, and an antiserum capable of neutralizing the cytotoxic effect of lymphotoxin was obtained. The anti-lymphotoxin activity was shown to be due predominantly to IgM immunoglobulin. The antiserum neutralized concanavalin A-induced lymphotoxin with equal efficiency as antigen-induced lymphotoxin. On the other hand, the anti-lymphotoxin serum did not neutralize preformed mitogenic factor or migration inhibitory factor (MIF) which had been produced by antigen-stimulated immune lymph node cells. Thus these activities appear to be due to substances which are antigenically distinct from lymphotoxin. Likewise, anti-lymphotoxin serum, when present in cultures of antigen-stimulated immune lymphoid cells, caused no diminution in the amount of mitogenic factor or MIF which was produced, indicating that it had no direct deleterious effect on the function of the lymphoid cells responsible for producing these lymphokines.


Modulation of the immune response by antigen-reactive lymphocytes after cultivation with gangliosides. GM1 ganglioside isolated from mouse brain was found to be capable of inhibiting the thymocytotoxicity of AKR anti-Thy 1.2 antiserum and to a lesser extent anti-BAtheta antiserum. GD1b ganglioside of mouse brain and thymocytes inhibited the thymocytotoxicity of rabbit anti-BAtheta anti-serum for mouse thymocytes. When GM1 ganglioside, in the form of cholesterol-lecithin liposomes, was incubated with spleen cell cultures in the presence of SRBC, depressed anti-SRBC hemolytic plaque responses were observed. This effect could be neutralized by first absorbing the GM1 liposomes with anti-Thy-1.2 antibodies. The kinetics of GM1 numbers when the ganglioside was added even as late as the 4th day of culture. Results of experiments in which GM1 ganglioside was incubated with either bone marrow cells or with thymocytes suggest that the target cell for GM1 liposomes is the B lymphocytes.


Differentiating feline T and B lymphocytes by rosette formation. The number of bone marrow-derived and thymus-dependent lymphocytes in the peripheral blood of felines was assessed. Feline bone marrow-derived lymphocytes were determined by erythrocyte antibody complement (EAC)-rosette formation and membrane immunofluorescence. A good correlation was observed between the cells forming EAC-rosettes (mean of 40.2 +/- 9.7%) and those possessing membrane immunoglobulins (mean of 45.3 +/- 4.1%). Feline thymus-dependent lymphocytes form non-immune (E)-rosettes with rodent erythrocytes (mouse, rat, guinea pig) but not with erythrocytes from humans, monkeys, dogs, mongooses, chickens, rabbits, sheep, and hamsters. An average of 32.3 +/- 992% of the peripheral blood lymphobytes of cats formed rosettes with guinea pig erythrocytes. Antibody blocking, cytotoxicity, and mix-rosette studies demonstrated that guinea pig red blood cells bind specifically to T lymphocytes. Of feline peripheral blood lymphocytes, 41.0 +/- 7.5% showed fluorescent staining with fluorescein-labeled rabbit anti-cat thymus antiserum.


Evidence that uridine incorporation is not a selective marker for mouse lymphocyte subclasses. We demonstrate here that mouse T+ and Ig+ lymphocytes do not show the differential incorporation of 3H-uridine into acid precipitable cellular components described for these lymphocyte subpopulation in the rat.


Investigation of the recovery phenomenon in immunofluorescence after laser excitation. Laser excitation was applied in two standard immunopathological indirect immunoflorescence systems in order to investigate bleaching and recovery of fluorescence intensity upon repeated illuminations. Variation of illumination times and dark periods between these showed that the recovery phenomenon is a function of both the (time times energy) product of excitation and the length of dark intervals. The value of high energy excitation in immunofluorescence in providing insight into the mechanisms of bleaching and recovery is discussed.


Cutaneous immunopathology: recent observations. Immunofluorescent studies are currently being done on patients with pemphigus, pemphigoid, dermatitis, lupus erythematosus and its variants, the cutaneous prophyrias, scarring alopecia, erosive mouth lesions, light-sensitive disorders, and cutaneous vasculitis. In this paper I shall review some of the recent advances in immunopathology and report the results that have been obtained in our laboratory.


The significance of cutaneous immunoglobulin deposits in lupus erythematosus and NZB/NZW F1 hybrid mice. In 50 to 60 percent of patients with systemic lupus erythematosus (SLE), a band of immunoglobulins beneath the epidermis of visibly normal skin. This highly specific finding has been used as a diagnostic test (Lupus Band Test) for SLE. Similar immunoglobulin deposits are found in an inbred strain of New Zealand mice which spontaneously develop an autoimmune disease with many features of SLE. Subepidermal immunoglobulin deposits are found most frequently in SLE patients with proliferative glomerulonephritis, hypocomplementemia, and serum antibodies to native DNA (anti-nDNA). When anti-nDNA levels are suppressed by cyclophosphamide, these deposits disappear. The subepidermal accumulation of immunoglobulin in SLE patients and in the mouse model apparently depends on the presence of antibody to native DNA. It is proposed that serum anti-nDNA precipitates with nDNA which is released locally from epidermal nuclear breakdown.


The epidermal-dermal junction.

The epidermal-dermal junction. Ultrastructurally, the epidermal-dermal junction is composed of four component areas:(1) the basal cell plasma membrane with its specialized attachment devices or hemidesmosomes, (2) an electron-lucent area, the lamina lucida, (3) the basal lamina, and (4) the sub-basal lamina fibrous components, including anchoring fibrils, dermal microfibril bundles, and collagen fibers. The light microscopic "basement membrane" comprises only the sub-basal lamina fibrous zone. Other cell types, including melanocytes and Merkel cells, are also found at the epidermal-dermal junction. Structures at the junction derive their origin from the epidermis and dermis: the basal lamina is primarily of epidermal origin, the anchoring fibrils of dermal origin. The junction serves the following functions: (1) epidermal-dermal adherence, (2) mechanical support for the epidermis, and (3) a barrier to the exchange of cells and of some large molecules across the junction.


Immunoglobulin classes of antibody to hepatitis B core antigen. Antibody to hepatitis B core antigen in the sera of 10 patients with acute viral hepatitis B, 20 patients with chronic hepatitis B, and 30 carriers of chronic hepatitis B surface antigen was assayed by the indirect immunofluorescence test. The activity of antibody to hepatitis B core antigen in the sera of these three groups of patients was identified as belonging to immunoglobulin classes G, M, A, and D. Antibody to hepatitis B core antigen was predominantly of IgG and IgM classes in all three groups of patients.


Spread of Streptococcus pneumoniae in families. I. Carriage rates and distribution of types. The mechanisms by which Streptococcus pneumoniae spreads from person to person are poorly understood. In this study, optimal methods for sampling, isolation, and identification of S. pneumoniae from healthy carriers were investigated. Factors influencing carriage rates were analyzed. Findings included the importance of pharyngeal rather than of nasal sampling in adults, the greater sensitivity of mouse inoculation compared with direct streaking of blood agar plates, and the superiority of the Quellung reaction with omniserum over the optochin disk as a means of identification of S. pneumoniae. Carriage rates were highest in children of preschool (35%) or grammar school (29%) age and in adults with preschool children in the family (18%). Rates were lowest in adults without preschool children in the family (2%-9%).


Synergism of amphotericin B and 5-fluorocytosine for candida species. The in vitro activity of amphotericin B and 5-fluorocytosine against 40 isolates of different species of Candida was studied. Synergism was seen in 35 of 40 organisms studied (88%) with achievable serum concentrations of the antibiotics. Synergism was striking in many strains that were resistant to 5-fluorocytosine. When the minimal inhibitory concentration of 5-fluorocytosine was greater than 400 mug/ml, synergism was seen in 18 of 20 isolates of Candida (90%).


New knowledge of chlamydiae and the diseases they cause. The trachoma and LGV organisms, the human pathogens of the species C. trachomatis, cause oculogenital infections and disease syndromes of the eye and genital tract. The incidence of the most prominent disease, endemic trachoma with eye-to-eye transmission, is decreasing all over the world. At the same time there is increasing recognition of high-frequency venereal infections with trachoma organisms and of the genital disease and occasional ocular disease that they cause. Laboratory techniques for diagnosis and investigation are improving, but work with these interesting intermediate agents remains more difficult than that with many other microorganisms. Proper recognition of the diseases is important because specific therapy is available.


Differential pulmonary residual volume determinations with radioactive xenon. A radioactive xenon inhalation procedure was used to determine differential residual volumes (right lung vs. left lung) in 28 normal subjects (6 of whom smoked approximately 1 package of cigarettes per day).  The average right lung residual volume represented 54% of the total residual volume, whereas the average left lung residual volumerepresented 46% of the total.  In addition, all of the subjects were studied by a standard helium dilution technique and found to have total lung residual comparable to the total lung residual volume as determined by the xenon technique.  There was no statistical difference between the data obtained from cigarette smokers vs. nonsmokers.  The noninvasive nature of the xenon technique makes this a useful procedure in determining differential residual volumes.


Incidence and titers of antinuclear, antismooth muscle, and other autoantibodies in blood donors. Sera from 107 blood donors ranging in age from 18 to 66 years were tested by indirect immunofluorescence for titers of various autoantibodies. These were matched for sex in diffferent age brackets. Antinuclear antibody (ANA) titiers of 10 or more appeared in 45 per cent of the sera examined. However, only 5.6 per cent had ANA titers of 80 or more, and only one subject had a titer of 160. ANA titers of 80 or greater were only found in the oldest age group (51 years or older). In those subjects with detectable ANA, 57.7 per cent were female and 32.8 per cent were male. The prevalence of thyroid cytoplasm and gastric parietal cell antibodies also appeared both age and sex related, while smooth muscle antibodies did not. Anti-DNA antibody, pemphigus antibody, bullous pemphigoid antibody, and mitochondrial antibody were detected in none of the sera tested. No relationships were seen in pairs of autoantibodies.


Chemotaxis as a preparative technique for human polymorphonuclear leukocytes. Human peripheral blood phagocytic cells were prepared in large quantities utilizing their chemotactic characteristics. Cells were isolated using an enlarged version of the Boyden chemotactic chamber and were collected after migrating through a Millipore membrane into a solution containing chemotactic factor. Chemotactic factors consisted of normal human serum, casein-saturated medium, and a combination of human serum and casein-saturated medium. The most effective attractant was the mixture of casein and fresh human serum. Yields of up to 24.8 times 10(6) cells could be attained from a starting volume of approximately 12 c.c. of peripheral blood. These cells were greater than 95 per cent polymorphonuclear leukocytes with the remaining cells being monocytes. No erythrocytes or lymphocytes contaminated these preparations. The advantages of this technique are as follows: (1) large quantities of human peripheral blood phagocytes consisting of greater than 95 per cent polymorphonuclear cells can be obtained: (2) these cells are known to be biologically active sice chemotaxis is a requirement for their isolation; (3) the separative method is dependent on chemotactic properties rather than sedimentation characteristics; and finally (4) the resulting cell preparations are virtually devoid of lymphocytes and erythrocytes.


Radioimmunoassay, acetylating radio-enzymatic assay, and microbioassay of gentamicin: a comparative study. Gentamicin is an aminoglycoside antibiotic widely used to treat gram-negative bacillary infections. Because it has a low therapeutic index, monitoring of serum levels may help to insure adequacy of dosage and avoid toxicity. Microbiological assays are relatively slow and can be complicated by the presence of other antimicrobials. Radioimmunoassay (RIA) and acetylating radio-enzymatic assay (ARA) are new methods for gentamicin assay which offer the following advantages: rapidity (less than 3 hours); no interference by other antibiotics; RIA is extremely sensitive and ARA is versatile (being useful in the measurement of other aminoglycosides). Correlation coefficients determined by linear regression analysis of assays on 36 patient samples performed in duplicate on 2 different days demonstrated no significant difference in measurement of gentamicin by each of the methods. Factors such as numbers of specimens, cost, and time involved will affect the decision of the method to be applied in individual laboratories.


Histopathological findings in cochlear implants in cats. This histopathological study on cats has shown that electrodes may be implanted into the cochlea through the round window or an opening drilled over the middle and apical turns without loss of the spiral ganglion cells or auditory nerve fibres. It has also shown that infection, leading to a labyrinthitis, was more likely to occur when an opening was drilled in the cochlea, and this could lead to a marked destruction of the cochlear structures, and loss of the spiral ganglion cells.


Leprosy of the upper respiratory tract. A clinical bacteriological, histopathological and histochemical study of twenty cases. Twenty cases clinically diagnosed as leprosy were thoroughly examined for E.N.T. lesions. These lesions were subjected to bacteriological, histopathological and histochemical studies. The results have been tabulated and discussed with special stress on some findings which are of help in diagnosing the disease.


P wave abnormalities in the orthogonal electrocardiogram: Correlation with ventricular overload in pulmonic and aortic valvular heart disease. The correlation between several P wave measurements form the orthogonal electrocardiogram (SVEC III lead system) extracted by computer analysis and simple hemodynamic parameters related to ventricular dysfunction was studied in two groups of patients. Group I consisted of 32 patients with pulmonic valvular stenosis and intact interventricular septum. There was a significant correlation between electrocardiographic criteria of right atrial overload and the two hemodynamic parameters studied: peak pulmonic systolic pressure gradient and right ventricular end diastolic pressure (r = 0.502, p smaller than 0.005 and r = 0.661, p smaller than 0.001 respectively). Group II consisted of 49 patients with aortic valve disease. In this group, a significant correlation between the electrocardiographic parameters of left atrial overload and the left ventricular end diastolic pressure could be demonstrated only be a multivariate regression analysis (r = 0.630, p smaller than 0.005). The P wave measurements that are well correlated with the ventricular end diastolic pressure can be considered as valuable criteria for atrial enlargement secondary to a decrease of ventricular compliance, such as seen in ventricular hypertrophy, failure or in ventricular constrictive or restrictive diseases. The pathophysiologic mechanisms of the influence of the ventricular overload (dysfunction) on the atrial function and the resulting P wave changes are discussed.


Semi-automated ECG processing: A simple method to improve efficiency in ECG laboratories. A method to expedite processing of electrocardiograms (ECGs) is described. The hardware configuration utilizes conventional equipment, and the ECG data is stored in magnetic data cards. The electrocardiographer's interpretation is made using a specially developed code of 253 diagnostic statements of 2 to 9 words each. A minicomputer converts the code into full alpha-numeric description and 2 characters into English statements. The diagnostic print-out appears in the same page as the reproduction of the original ECG data. This system has significantly reduced the ECG processing time, freed manpower to increase availability of technicians and decreased the size of permanent files.


Quantitative vectorcardiographic criteria for the differentiation between atrial septal defects of primum and secundum types. From a population of 304 patients who were operated on for atrial septal defects, 25 patients with primum defects and 78 patients with secundum defects were studied with regard to the ability of various quantitative axial lead vectorcardiographic (VCG) data to predict the anatomical type of defect. The patients included all in whom there had been preoperative doubt concerning the classification. By applying the electrocardiographic (ECG) mean frontal QRS axis, 8 patients were erroneously classified, and 14 could not be classified because no definite QRS resultant was present in the frontal plane (diagnostic performance index 0.85). Several VCG criteria showed a better performance, the best being sums of 10 msec Y lead amplitudes from 20 to 50 msec after QRS onset. These criteria, which reflect the superior shift of the middle part of the QRS complex in primum defects, reduced the number of misclassifications to a minimum of 4 (performance 0.96). A combination of this criterion with one based on the duration of the initial inferior QRS vectors gave some further improvement. This simple combination was not surpassed by classification performed by a stepwise discriminant analysis computer program (BMD 07M).


Electrocardiographic screening by special purpose computer. Correlation with twelve-lead electrocardiogram. An electrocardiometer (ECM) which computes the T/R ratio of Lead I and simultaneously prints out Lead I was used to screen 1,004 consecutive hospitalized patients for EKG abnormalities. When the combined meter and Lead I print out analyses were used, 77.9% of abnormal EKG's were identified with a false-positive rate of less than 4%. The method is highly specific for detecting EKG abnormalities associated with anterior and lateral left ventricular disease. It could be used as an inexpensive mass screening device for electrocardiographic risk factors associated with coronary disease.


The responsiveness of the anterior pituitary gland to luteinizing hormone releasing factor in rats exposed to constant light. The responsiveness of the anterior pituitary gland to synthetic luteinizing hormone releasing factor (LH-RF) was tested in rats exposed to constant light. At a dosage of 50 ng LH-RF/100 g body wt the mean maximal increments in plasma LH and FSH were similar to those at 10.00 h of pro-oestrus. The increments in the plasma gonadotrophins at dosages of 500 and 1000 ng LH-RF/100 g body wt did not differ significantly from those at 250 ng LH-RF/100 g body wt. These findings suggest that, in contrast to rats which exhibit regular oestrous cycles, the preovulatory (post-coital) release of LH in rats exposed to constant light may depend almost entirely on the release of a relatively large amount of LH-RF into hypophysial portal vessel blood. Whereas in pro-oestrous animals a relatively small fraction of the readily releasable pool of LH is released during the spontaneous preovulatory surge, in rats exposed to constant light most releasable LH appears to be discharged during the reflex preovulatory surge of this hormone. The concentrations of radioimmunoassayable FSH in blood samples withdrawn before the injection of LH-RF support the view that FSH secretion in the rat is increased by constant exposure to light.


The effects of minute direct electrical currents on cultured chick embryo trigeminal ganglia. The effects of low levels of electric current were determined on organ cultures of 6- to 19-day-old chick embryo trigeminal ganglia. Current levels ranged from 0.00115 to 11.5 nA/mm2; the time of electrical treatment varied from 2 h to 96 h. Low levels of electricity were found to have at least three major effects on this system of mixed cellular types: (1) Outgrowth of nerve fibers from the explant was enriched. This resulted in a greater number of fibers which were longer and more highly branched than those in control cultures. (2) Survival of neurons within the original explanted ganglion was enchanced by treatment with electricity. The presence of healthy neurons was found in 93.1% of the treated cultures and in 53.5% of untreated cultures. (3) Neurons, fibers and non-neural cells were stimulated to grow in the direction of the cathode. The rate of cathodal migration was calculated to be 0.1 mm/h (2.4 mm/day) in an explant from 12-day-old embryo. No differences were observed between surviving treated and control cultures in histochemical tests for acetycholinesterase. A similarity between the action of low levels of electrical current and nerve growth factor (NGF) is suggested.


Histocompatibility studies in a closely bred colony of dogs. V. Mechanisms of cellular adaptation in long-term DL-A identical radiation chimeras. 20 Cooperstown beagles of known DL-A genotypes (B" dogs) were exposed to supralethal total body irradiation and received a bone marrow allograft from a DL-A identical donor (A" dog); the resulting chimeras have survived uneventfully for 882, 1466 days, with no evidence of secondary disease, and have been tolerant to kidney and skin allografts obtained from the donor of marrow. Treatment of "A" dogs with serum obtained from their long-term "B" chimeras had no significant effect upon the ability of the recipients to reject "B" skin allografts...


C1q deviation test for the detection of immune complexes, aggregates of IgG, and bacterial products in human serum. This report describes a new, rapid, sensitive, and quantitative method for the detection of immune complexes, endotoxins, and other complement activating materials in patients sera utilizing the ability of these substances to react with isolated C1q. The procedure is based on the inhibition of radiolabeled C1q binding to sensitized sheep erythrocytes by C1q-reactive substances in pathological sera. The C1q deviation test may be performed on 50 mu1 of serum, using 1 mug of radiolabeled C1q per sample. The procedure may be completed in 1.5-2 h, it is capable of detecting 5 mug of aggregated human IgG per ml of serum, and its coefficient of variation is 4.2%. Application of the test to the study of 193 sera from 43 patients with Dengue hemorrhagic fever showed a positive correlation between degree of C1q deviation and severity of disease.


The modulation of lymphocyte functions by molecules secreted by macrophages. I. Description and partial biochemical analysis. Culture fluids of peritoneal exudate cells rich in macrophages stimulated DNA synthesis of thymocytes and, to lesser extent, of spleen cells. We also investigated the effects of culture fluids from macrophages on the in vitro response to a hapten-carrier protein (fluorescein-menocyanin) using spleen cells from immune mice. Macrophage culture fluids contained an activity that increased the plaque-forming cell response of both IgG and IgM class. This increase was observed in the absence of any added hapten protein to the culture. The helper function of T lymphocytes (as evidenced by challenging with the hapten on the homologous carrier) was also increased by the macrophage culture fluid. However, this enhancement was best observed in conditions of relatively low T-cell activity. Also, the macrophage fluid allowed spleen cells of nude athymic mice to make a plaque-forming cell response to sheep red blood cells of both the IgM and IgG class. The macrophage was the cell source of the stimulatory molecule since it was generated only in cultures of macrophages devoid of significant number of lymphocytes. Stimulatory activity was not found in cultures of lymphocytes, mouse embryo cells, or 3T3 cells. The thymocyte stimulatory molecule did not contain H-2 antigens, was resistant to diisopropylfluorophosphate treatment, eluted from Sephadex with a size ranging from 15,000 to 21,000 daltons, and was sensitive to chymotrypsin and pepsin.


Multiple mouse-protective antibodies directed against group B streptococci. Special reference to antibodies effective against protein antigens. The data presented in this paper establish the finding that multiple specific protective antibodies exist in rabbits in response to immunization with Group B streptococci. The summary in Table I indicates the serological types into which Group B streptococci have been divided on the basis of their antigenic composition. This classification is dependent upon passive protection of mice with antibodies directed against the specific antigens, and types are defined in these terms. Heretofore, it was thought that type-specific polysaccharides accounted for all such protection in Group B streptococci. Certain exceptions of cross-protection between types due to minor polysaccharide determinants soon appeared; cross-protection reactions based on protein determinants in at least two types were also discovered. The present experiments show that specific antibodies directed to either polysaccharide or protein antigens of a single strain can be protective against infection with streptococci containing these antigens.