in this retrospective , single center study , we reviewed charts from 68 patients with ana comprehensive panels .
inclusion criteria were patients 18 years old and had an ana cp billing code between may 2015 and october 2015 .
variables included age , sex , specialty of the ordering physician , test indication , and ana result .
our institutional review board determined this study was non - human subjects research ; institutional approval was not required .
the mean age of the sample was 54.419.4 years old , and 60.3% were female .
internal medicine ordered the majority of ana cps ( 83.8% ) followed by family medicine ( 7.4% ) , emergency medicine ( 2.2% ) , and psychiatry ( 2.2% ) .
hypercoagulable work up , transaminasemia and skin rash were the most frequent indications for ordering the ana cp ( 8.8% for each indication ) .
the remaining indications ( 73.6% ) covered a broad spectrum and combined as an  other category .
all the ana cps ordered were considered to be inappropriate including the three patients who had previous history of rheumatological disease and did not require re - testing .
sixty - three ana comprehensive panels were negative for rheumatological disease ( 92.6% , fig .
multiplex immunoassays have led to a paradigm shift in the methodological testing of autoimmune diseases .
high throughput multiplex immunoassays have supplanted the use of traditional methods like indirect immunofluorescence ( iif ) and elisa .
iif testing is subject to poor specificity , has a high false positive rate , lack of standardization in substrate and dilution protocols , and interobserver variability in pattern interpretation ( 2 ) . enzyme immunoassays ( eia ) testing removes the subjective variations of iif testing ( 3 ) ; however , there exist interlaboratory method variations and heterophile antibody interferences causing false - positive results .
the correlation between elisa and mutiplex assays is high , with a 90% concordance ( 4 ) .
ana testing with multiplexed microsphere fluorescence allows for rapid quantification and efficient profiling of multiple clinically significant antibodies in a single run of assay ( 5 ) .
the multiplex ana screen is a composite screen which tests for 11 specific autoantibodies that are known to be associated with autoimmune diseases .
if none of the specific antibodies are present , the ana screen is reported as negative .
positive screens are reflexed , and the reflexed antibodies are resulted semi - quantitatively as numeric antibody indices ( ai ) ( 5 ) . the authors identified that the major reason behind inappropriate 
ana comprehensive panel ordering could be due to physician unawareness regarding test components performed under the order panel .
ana testing by mutiplex assays should be performed in a bi - leveled sequence with an initial screen followed by comprehensive antibody testing only if ana screen returns positive .
ana comprehensive panel is not a reflex laboratory test and examines directly for specific antibodies without going through an initial screen .
comprehensive antibody testing without an initial screen leads to unnecessary cost burden on the patient and a waste of laboratory resources . in our study ,
net calculated value of incurred losses resulting from  comprehensive lab order ( ana cp ) testing amounted to $ 66,000 over a 6-month study period .
the  ana comprehensive panel does not provide any further utility over the  ana screen with reflex. hence , the ideal test of choice should be  ana screen with reflex. also , the term ana comprehensive panel gives the physicians a sense of it being a more complete test over the ana screen , thus leading to inappropriate use of the second - level testing without going through the initial screen . over the last few years , ana comprehensive panels have emerged as a very convenient test with fewer false positives and the convenience of being completed in a single run , resulting in the ordering physician choosing it over the appropriate ana screen .
it is important to recognize that autoantibody testing is fraught with low specificity and false - positive results , leading to potential over - diagnosis and mistreatment .
in fact , false positives occur in a significant proportion of healthy population ( 6 ) .
ana testing in the wrong clinical setting , especially in the absence of specific signs and symptoms suggestive of rheumatic diseases can potentially lead to misdiagnosis and unnecessary rheumatology consultations . also , repeat ana testing is not warranted in patients with established rheumatological disease ( as noted in 3 of 68 in our study patients ) except to monitor disease activity , in which case , repeating specific antibody titers may be considered and not the  ana comprehensive panel. a retrospective study on patients who had ana testing suggested that a large proportion ( 30% ) of them had non - specific and unrelated signs and symptoms ( 7 ) .
incorporation of ana testing into initial lab order bundles in an effort to expedite diagnoses ; overestimation of the pretest probabilities ; failure to consider non - rheumatological factors expected to cause false - positive results contribute to ana testing among physicians .
one of the limitations of our study was that it was a single institutional experience .
we initially conducted this as a pilot study and based on the results in our center , we intend to conduct the study across all advocate healthcare hospitals .
ana testing with multiplex immunoassay systems allows for rapid profiling of multiple analytes in a single run of assay and have few false - positive results compared to conventional lab methods .
however , it is important to educate physicians on following a sequenced diagnostic algorithm based on the guidelines laid down by american college of rheumatology to avoid unnecessary higher level testing and increase cost burden on their patients .
also , inappropriate ana testing without adequate pretest probability estimation after careful history taking and clinical examination can lead to over - diagnosis with unnecessary consultations and should be avoided .
the authors have not received any funding or benefits from industry or elsewhere to conduct this study
 healthcare providers use antinuclear antibodies ( anas ) to screen and diagnose patients with autoimmune diseases . in the recent years , commercial multiplex ana kits have emerged as a convenient and fast diagnostic method . 
 diagnostic testing should follow sequenced algorithms : initial screen followed by specific antibody analysis . 
 second - level testing as an initial screen for autoimmune disease is inappropriate . 
 we reviewed 68 patients with ana comprehensive panels over a 6-month period from may 2015 to october 2015 . 
 we assessed appropriateness and estimated incurred losses from inappropriate testing . 
 we found 92.6% ( 63 out of 68 ) of the ana comprehensive panel results to be negative . 
 incurred losses from inappropriate ana comprehensive panel testing were $ 66,000 . 
 physicians should become familiar with ana - sequenced diagnostic algorithms to avoid unnecessary higher level testing . 
Methods
Results
Discussion
Conclusion
Conflict of interest and funding