Detection of HLA-specific IgG antibodies using single recombinant HLA alleles - The monoLISA assay

Background. Because of the presence of confounding antigens, the assignment of HLA antibody specificity is difficult; in highly sensitized patients, a nd the definition of an acceptable HLA mismatch requires a significant work load per patient. We describe a new ELISA method, monoLISA, for detection o f immunoglobulin (Ig)G HLA antibody using single recombinant HLA class I mo nomers bound to microtiter plates. Methods, HLA-A2 and -B8 monomers were synthesized and used as screening tar gets for 85 sera from renal patients. The sera contained various IgG and Ig M HLA-specific antibodies, including anti-A2 and anti-B8,defined in a conve ntional complement-dependent cytotoxicity test (CDC). Investigations were p erformed to determine possible effects on antibody binding of differential monomer peptide presentation as well as lack of glycosylation, Results. A good correlation was found between CDC-defined specificities and the reactivity observed with HLA monomers, MonoLISA attained means of 100% sensitivity and 92.5% specificity compared with CDC, Neither the presence of different peptides, nor the absence of glycosylation of the monomer affe cted the ability of monoLISA to detect antibody. Conclusion. This study demonstrates that the monoLISA method for HLA antibo dy detection is valid. Because this has the potential to reduce the work in volved in screening sensitized patients awaiting transplantation for HLA an tibodies, resources aimed at increasing the number of constructed monomers would be well targeted.