Patients who have become sensitized to allogeneic HLA antigens through pregnancies, blood transfusions, failed transplants, or other means represent a challenge when requiring a transplant or platelet transfusion. Preformed anti-HLA antibodies are associated with hyperacute rejection of kidney, heart and lung grafts, primary nonfunction of liver grafts, failure of platelet transfusions and accelerated and chronic loss of solid organ transplants. When present in blood or plasma donors, preformed anti-HLA antibodies may cause transfusion-related acute lung injury (TRALI) in some recipients. These antibodies can be identified and characterized using several approaches which vary in sensitivity and accuracy.
Tests for the present of circulating anti-HLA are critical for:
Blood and platelet transfusions
Blood and plasma donors
Anti-HLA antibodies in patient's serum may react with cells from a few or many individuals. Sensitization is reported as the percent Panel Reactive Antibody (PRA), which is the percentage of potential donors' cells tested that were killed by the patient's serum. The PRA also gives an estimate of the likelihood of finding a suitable donor. Computerized analyses of lymphocytotoxicity reaction patterns on well-characterized cell panels identify the specific HLA-antigens against which antibodies are directed, and "safe" antigens, which are not recognized. Patients who have autoimmune diseases, are on particular medications, or have infections may develop false positive reactions that are not caused by anti-HLA antibodies. Flow bead tests provide results separately for class I and class II antibodies and since they are based on purified HLA antigens, they avoid many of the false positive reactions. Flow bead testing can also be used to identify HLA specificities even with low antibody titers.
Results are reported as the percent PRA for cytotoxicity tests, whether or not reactivity is DTT sensitive, and the anti-HLA specificities that were identified. Luminex and flow cytometry test reports indicate percent PRA for class I and class II antigens and antibody specificity. Single recombinant HLA class I and class II antigens coupled to microparticles are used to determine the HLA specificity of broadly reactive sera or sera with undefined specificity.
Anti-HLA antibody testing provides an accurate assessment of an individuals sensitization status and identification of the HLA antigens specifically targeted by those antibodies. The tests used to identify anti-HLA antibodies are based upon assessing reactions of antibodies with well-characterized panels of individual lymphocytes (using complement-dependent lymphocytotoxicity), and with purified HLA antigens coupled to specialized microparticles (luminex, flow cytometry).
Complement-dependent lymphocytotoxicity is used to identify patients at risk for antibody-mediated damage to grafted organs or tissue. This test can also be performed after transplantation.
Luminex is used before and after transplantation to monitor patients for presence or absence of anti-HLA class I and class II antibodies or to monitor the appearance or disappearance of antibodies over time. Positive sera are subsequently tested by luminex class I and class II identification assays to determine the HLA specificity of the antibodies.
Flow cytometry is used before and after transplantation to assess the specificity of anti-HLA antibodies that are not normally detected by less sensitive methods such as cytotoxicity. Single HLA antigen beads are used to determine specific HLA antibodies in high PRA containing sera.
The MICA antibody test detects antibodies against a panel of MICA single antigens. The test detects antibodies directed against the most common MICA alleles and allows high throughput antibody screening and identification using luminex technology. The test can be performed before or after transplantation.