Determination of the presence of an antigen, particularly HLA-B27 antigen, at the surface of a cell, particularly lymphocytes, by cytotoxicity test combined with fluorescent staining

摘要

The present invention relates to a method of determining the presence of an antigen, particularly the HLA-B.sub.27 antigen, at the surface of cells, particularly lymphocytes, by combining the sensitivity of a cytotoxicity test, particularly a lymphotoxicity test, combined to a fluororescence detection, particularly, by using propidium iodide. The claimed method does not involve any purification of lymphocytes from a blood sample. The erythrocytes contained in the test blood sample are lysed after the immune reaction and complement action is completed. Moreover, the formation of the immune complexes, the complement fixation and reaction and the staining of the cells may take place at the same time in the test tube. This method is therefore very simple and very fast to execute. The facility of reading the positive cells is also an advantage to the claimed method over and above the methods using dyes like eosin or trypan blue, because of the direct visualization of colored cells as positive cells. Another advantage of the claimed method resides in its versatility of reading. Quantitation and identification of the positive cells is rendered feasible by fluorescence microscopy as well as by flow cytofluorometry. The claimed method has been validated in the diagnosis of ankylosing spondylitis (AS) and reactive arthritis (RA) by typing lymphocytes HLA-B.sub.27.