HLA-B27 determination using serological methods. A comparison of enzyme immunoassay and a microlymphocytotoxic test with flow cytometry and a molecular biological assay.
Typing for HLA-B27 is routinely performed in patients with seronegative spondarthritides. Besides the microlymphocytotoxic test (MLCT), other serological techniques have been developed such as enzyme immunoassays (EIA) using serum or plasma as a source for the determination of soluble HLA-B27 (sHLA-B27) and flow cytometric (FC) methods. The aim of the present study was to check the accuracy and reliability of the EIA for sHLA-B27 in comparison to the MLCT using antibodies against HLA-B27 and cross-reacting specificities (CRS), as well as an FC method and a molecular biological method. Any discrepant results should be typed with the MLCT using a complete panel of anti-HLA-class I antibodies, with FC and with a molecular biological technique. The EIA should also be repeated in those patients, using serum and plasma from a new venipuncture. In 81 patients with rheumatic disorders, the EIA and the MLCT using antibodies against HLA-B27 and CRS were performed. Based on the MLCT with a complete panel of anti-HLA-class I antibodies as a standard, discrepant test results were obtained for 9 out of 81 patients with the MLCT using antibodies against HLA-B27 and CRS and with the EIA. The following wrong results occurred: in the MLCT with anti-HLA-B27 and CRS, there were two false-negative results; in the EIA there were four false-negative and one false-positive results; one sample was undeterminable. In comparison with the MLCT, including the complete panel of HLA-class I antibodies, as well as with a molecular biological technique, typing with FC showed a complete concordance. Our investigations demonstrated that for routine typing for HLA-B27 the MLCT cannot be replaced by EIA because of a significant number of mistypings. The MLCT performed only with antibodies against HLA-B27 and CRS may also lead to typing errors. No errors were detected using flow cytometry. If only serological methods can be performed in a laboratory a combination of flow cytometry and MLCT could therefore enhance the safety of HLA-B27 typing.
5th Department of Internal Medicine, Wilhelminenspital, Vienna, Austria.
A
A
Dunky
J
J
Neumüller
C
C
Hübner
G F
GF
Fischer
P M
PM
Bayer
E
E
Wagner
D W
DW
Schwartz
W R
WR
Mayr
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000911:Antibodies, Monoclonal / Q000032:analysis; D003601:Cytotoxicity Tests, Immunologic; D017931:DNA Primers; D005260:Female; D005434:Flow Cytometry; D015796:HLA-B27 Antigen / Q000097:blood* / Q000235:genetics / Q000276:immunology; D006801:Humans; D007124:Immunoenzyme Techniques; D008297:Male; D008875:Middle Aged; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length
D000911:Antibodies, Monoclonal; D017931:DNA Primers; D015796:HLA-B27 Antigen