Systematic comparison of four cell- and Luminex-based methods for assessment of complement-activating HLA antibodies.
BACKGROUND
Efforts to increase the specificity and sensitivity of human leukocyte antigen (HLA) antibody detection assays recently led to the establishment of two novel Luminex bead-based assays to detect complement-activating antibodies by the assessment of complement products C1q or C4d. Here, we present a systematic comparison of the four methods, complement-dependent lymphocytotoxicity (CDC) and C1q-, C4d-, and IgG-Luminex, to assess or predict the complement-binding capability of HLA IgG antibodies.


METHODS
Forty-five sera of highly immunized patients have been assessed by in-house modified C1q- and C4d-Luminex assays and compared with standard CDC and IgG-Luminex.


RESULTS
Antibody specificities assigned by the C1q- and C4d-Luminex assay revealed an excellent concordance of 94% and 97% for HLA class I and II, respectively. Complement-fixing HLA class II antibodies were found less frequently among IgG antibodies compared with class I. Both C1q- and C4d-Luminex detected, on average, three times more specificities than CDC. Although we found a high correlation of mean fluorescence intensity values between C1q- and C4d-Luminex assays, IgG mean fluorescence intensity was not a suitable surrogate marker for the prediction of complement binding.


CONCLUSIONS
C1q- and C4d-Luminex assays are characterized by an increased sensitivity and specificity compared with CDC, the current standard in detecting complement-fixing HLA antibodies. Pretransplantation risk assessment for transplantation but also posttransplantation monitoring are important applications for both assays to improve overall allograft survival.
Charité-Universitätsmedizin Berlin, Center for Tumor Medicine and ZTB Zentrum für Transfusionsmedizin und Zelltherapie Berlin, Tissue Typing Laboratory, Berlin, Germany.
Nils
N
Lachmann
Kremena
K
Todorova
Harald
H
Schulze
Constanze
C
Schönemann
D015922:Complement C1q / Q000276:immunology; D015935:Complement C4b / Q000276:immunology; D003165:Complement System Proteins / Q000276:immunology*; D003602:Cytotoxicity, Immunologic; D006680:HLA Antigens / Q000276:immunology; D006801:Humans; D007114:Immunization; D007118:Immunoassay / Q000379:methods*; D007518:Isoantibodies / Q000097:blood*; D010446:Peptide Fragments / Q000276:immunology
D006680:HLA Antigens; D007518:Isoantibodies; D010446:Peptide Fragments; D015922:Complement C1q; D015935:Complement C4b; C032261:complement C4d; D003165:Complement System Proteins