reproductive organs. pap smears may trigger gender dysphoria in patients and gender-neutral language can be used when explaining the pathogenesis of cancer due to infection, emphasizing the pervasiveness of hpv infection regardless of gender. procedure cervix in relation to upper part of vagina and posterior portion of uterus. transformation zone types, determining the pap test location:type 1: completely ectocervical.type 2: endocervical component but fully visible.type 3: endocervical component, not fully visible. for best results, a pap test should not occur when a woman is menstruating, partly because the additional cells can obscure cervical cells and partly because this part of the menstrual cycle is when the female organs are most inflamed. however, pap smears can be performed during a woman's menstrual period, especially if the physician is using a liquid-based test; if bleeding is extremely heavy, endometrial cells can obscure cervical cells, and it is therefore inadvisable to have a pap smear if bleeding is excessive. obtaining a pap smear should not cause much pain, but it can if the woman has certain untreated vaginal problems such as cervical stenosis or vaginismus. it is, however, not comfortable, for two reasons: the cervix is full of nociceptors (pain nerves), and the brush used to collect cells has to be stiff enough to scrape them off of the surrounding tissue. so it can be uncomfortable, but it is generally quick, and the information obtained may be critical. people with underlying pain or tissue diseases that can react to nociceptors being scraped or to excessive cold in the mucous membranes should take appropriate precautions and discuss the process ahead of time with their providers. many people experience spotting or mild diarrhea afterward. the spotting is usually from the scrape on the cervix, and the diarrhea may be due to indirect stimulation of the lower intestine during the exam. a number of studies have shown that using a small amount of water-based gel lubricant does not interfere with, obscure, or distort the pap smear. further, cytology is not affected, nor are some std testing. the health care worker begins by inserting a speculum into the woman's vagina, which spreads the vagina open and allows access to the cervix. the health care provider then collects a sample of cells from the outer opening or os of the cervix by scraping it with an aylesbury spatula. an endocervical brush is rotated in the central opening of the cervix. the cells are placed on a glass slide and taken to the laboratory to be checked for abnormalities. a plastic-fronded broom is sometimes used in place of the spatula and brush. the broom is not as good a collection device, since it is much less effective at collecting endocervical material than the spatula and brush. the broom is used more frequently with the advent of liquid-based cytology, although either type of collection device may be used with either type of cytology. the sample is stained using the papanicolaou technique, in which tinctorial dyes and acids are selectively retained by cells. unstained cells cannot be seen adequately with a light microscope. papanicolaou chose stains that highlighted cytoplasmic keratinization, which actually has almost nothing to do with the nuclear features used to make diagnoses now. a single smear has an area of 25 x 50 mm and contains a few hundred thousand cells on average. screening with light microscopy is first done on low (10x) power and then switched to higher (40x) power upon viewing suspicious findings. cells are analyzed under high power for morphologic changes indicative of malignancy (including enlarged and irregularly shaped nucleus, an increase in nucleus to cytoplasm ratio, and more coarse and irregular chromatin). approximately 1,000 fields of view are required on 10x power for screening of a single sample, which takes on average 5 to 10 minutes. in some cases, a computer system may prescreen the slides, indicating those that do not need examination by a person or highlighting areas for special attention. the sample is then usually screened by a specially trained and qualified cytotechnologist using a light microscope. the terminology for who screens the sample varies according to the country; in the uk, the personnel are known as cytoscreeners, biomedical scientists (bms), advanced practitioners and pathologists. the latter two take responsibility for reporting the abnormal sample, which may require further investigation. automated analysis in the last decade, there have been successful attempts to develop automated, computer image analysis systems for screening. although, on the available evidence automated cervical screening could not be recommended for implementation into a national screening program, a recent nhs health technology appraisal concluded that the 'general case for automated image analysis ha(d) probably been made'. automation may improve sensitivity and reduce unsatisfactory specimens. two systems have been approved by the fda and function in high-volume reference laboratories, with human oversight. types of screening for other cervical screening tests and human papillomavirus testing, see cervical screening. conventional pap—in a conventional pap smear, samples are smeared directly onto a microscope slide after collection. however this method hurts as wooden spatulas are used. this causes pain during the procedure. liquid-based cytology—the sample of (epithelial) cells is taken from the transitional zone, the squamocolumnar junction of the cervix, between the ectocervix and the endocervix. liquid-based cytology often uses an arrow-shaped brush rather than the conventional spatula. the cells taken are suspended in a bottle of preservative for transport to the laboratory, where they are analyzed using pap stains. squamous metaplasia of the cervix, with typical features. pap stain. pap tests commonly look for epithelial abnormalities/ metaplasia/ dysplasia/ borderline changes, all of which may be indicative of cin. nuclei will stain dark blue, squamous cells will stain green and keratinised cells will stain pink/ orange. koilocytes may be observed where there is some dyskaryosis (of epithelium). the nucleus in koilocytes is typically irregular, indicating possible cause for concern; requiring further confirmatory screens and tests. in addition, human papillomavirus (hpv) test may be performed either as indicated for abnormal pap results, or in some cases, dual testing is done, where both a pap smear and an hpv test are done at the same time (also called pap co-testing). practical aspects the endocervix may be partially sampled with the device used to obtain the ectocervical sample, but due to the anatomy of this area, consistent and reliable sampling cannot be guaranteed. since abnormal endocervical cells may be sampled, those examining them are taught to recognize them. the endometrium is not directly sampled with the device used to sample the ectocervix. cells may exfoliate onto the cervix and be collected from there, so as with endocervical cells, abnormal cells can be recognised if present but the pap test should not be used as a screening tool for endometrial malignancy. in the united states, a pap test itself costs $20 to $30, but the costs for pap test visits can cost over $1,000, largely because additional tests are added that may or may not be necessary. history the test was invented by and named after the greek doctor georgios papanikolaou, who started his research in 1923. aurel babeș independently made similar discoveries in 1927. however, babeș' method was radically different from papanikolaou's. the pap test was finally recognized only after a leading article in the american journal of obstetrics and gynecology in 1941 by papanikolaou and herbert f. traut, an american gynecologist. a monograph entitled diagnosis of uterine cancer by the vaginal smear that they published contained drawings of the various cells seen in patients with no disease, inflammatory conditions, and preclinical and clinical carcinoma. the monograph was illustrated by hashime murayama, who later became a staff illustrator with the national geographic society. both papanikolaou and his wife, andromachi papanikolaou, dedicated the rest of their lives to teaching the technique to other physicians and laboratory personnel. experimental techniques in the developed world, cervical biopsy guided by colposcopy is considered the "gold standard" for diagnosing cervical abnormalities after an abnormal pap smear. other techniques such as triple smear are also done after an abnormal pap smear. the procedure requires a trained colposcopist and can be expensive to perform. however, pap smears are very sensitive and some negative biopsy results may represent undersampling of the lesion in the biopsy, so negative biopsy with positive cytology requires careful follow-up. experimental visualization techniques use broad-band light (e.g., direct visualization, speculoscopy, cervicography, visual inspection with acetic acid or with lugol's, and colposcopy) and electronic detection methods (e.g., polarprobe and in vivo spectroscopy). these techniques are less expensive and can be performed with significantly less training. they do not perform as well as pap smear screening and colposcopy. at this point, these techniques have not been validated by large-scale trials and are not in general use. access australia australia has used the pap test as part of its cervical screening program since its implementation in 1991 which required women past the age of 18 be tested every two years. in december 2017 australia discontinued its use of the pap test and replaced it with a new hpv test that is only required to be conducted once every five years from the age of 25. medicare covers the costs of testing; however, a patient's doctor does not allow bulk billing, they may have to pay for the appointment and then claim the medicare rebate. taiwan free pap tests were offered from 1974–1984 before being replaced by a system in which all women over the age of 30 could have the cost of their pap test reimbursed by the national health insurance in 1995. this policy was still ongoing in 2018 and encouraged women to screen at least every three years. despite this, the number of people receiving pap tests remain lower than countries like australia. some believe this is due to a lack of awareness regarding the test and its availability. it has also been found that women who have chronic diseases or other reproductive diseases are less likely to receive the test. england as of 2020 the nhs maintains a cervical screening program in which women between the age of 25–49 are invited for a smear test every three years, and women past 50 every five years. much like australia, england uses a hpv test before examining cells that test positive using the pap test. the test is free as part of the national cervical screening program. coccoid bacteria the finding of coccoid bacteria on a pap test is of no consequence with otherwise normal test findings and no infectious symptoms. however, if there is enough inflammation to obscure the detection of precancerous and cancerous processes, it may indicate treatment with a broad-spectrum antibiotic for streptococci and anaerobic bacteria (such as metronidazole and amoxicillin) before repeating the smear. alternatively, the test will be repeated at an earlier time than it would otherwise. if there are symptoms of vaginal discharge, bad odor or irritation, the presence of coccoid bacteria also may indicate treatment with antibiotics as per above.