Graft Immunocomplex Capture Fluorescence Analysis to Detect Donor-Specific Antibodies and HLA Antigen Complexes in the Allograft.
BACKGROUND
Immunocomplex capture fluorescence analysis (ICFA) is an attractive method to detect donor-specific anti-HLA antibodies (DSA) and HLA antigen complexes. Currently, antibody-mediated rejection (AMR) due to DSA is usually diagnosed by C4d deposition and serological DSA detection. Conversely, there is a discrepancy between these findings frequently. Thereupon, our graft ICFA technique may contribute to establish the diagnosis of AMR.


METHODS
Graft samples were obtained by a percutaneous needle biopsy. Then, the specimen was dissolved in PBS by the lysis buffer. Subsequently, HLA antigens were captured by anti-HLA beads. Then, DSA-HLA complexes were detected by PE-conjugated anti-human IgG antibodies, where DSA had already reacted with the allograft in vivo, analyzed by a Luminex system.


RESULTS
A ratio (sample MFI/blank beads MFI) was calculated: ≥ 1.0 was determined as positive. We found that DSA-HLA complexes in the graft were successfully detected from only slight positive 1.03 to 79.27 in a chronic active AMR patient by graft ICFA. Next, positive graft ICFA had predicted the early phase of AMR (MFI ratio: 1.38) even in patients with no serum DSA. Finally, appropriate therapies for AMR deleted DSA deposition (MFI ratio from 0.3 to 0.7) from allografts.


CONCLUSIONS
This novel application would detect early phase or incomplete pathological cases of AMR, which could lead to a correct diagnosis and initiation of appropriate therapies. Moreover, graft ICFA might address a variety of long-standing questions in terms of DSA.


BACKGROUND
AMR: Antibody-mediated rejection; DSA: Donor-specific antibodies; ICFA: Immunocomplex capture fluorescence analysis.
a Department of Organ Transplantation and General Surgery , Kyoto Prefectural University of Medicine , Kyoto , Japan.
Tsukasa
T
Nakamura
a Department of Organ Transplantation and General Surgery , Kyoto Prefectural University of Medicine , Kyoto , Japan.
Hidetaka
H
Ushigome
b Department of Blood Transfusion and Cell Therapy , Kyoto Prefectural University of Medicine , Kyoto , Japan.
Kiyoko
K
Watabe
b Department of Blood Transfusion and Cell Therapy , Kyoto Prefectural University of Medicine , Kyoto , Japan.
Yui
Y
Imanishi
a Department of Organ Transplantation and General Surgery , Kyoto Prefectural University of Medicine , Kyoto , Japan.
Koji
K
Masuda
a Department of Organ Transplantation and General Surgery , Kyoto Prefectural University of Medicine , Kyoto , Japan.
Takehisa
T
Matsuyama
a Department of Organ Transplantation and General Surgery , Kyoto Prefectural University of Medicine , Kyoto , Japan.
Shumpei
S
Harada
a Department of Organ Transplantation and General Surgery , Kyoto Prefectural University of Medicine , Kyoto , Japan.
Katsuhiro
K
Koshino
a Department of Organ Transplantation and General Surgery , Kyoto Prefectural University of Medicine , Kyoto , Japan.
Taku
T
Iida
a Department of Organ Transplantation and General Surgery , Kyoto Prefectural University of Medicine , Kyoto , Japan.
Shuji
S
Nobori
a Department of Organ Transplantation and General Surgery , Kyoto Prefectural University of Medicine , Kyoto , Japan.
Norio
N
Yoshimura
D000328:Adult; D000368:Aged; D064591:Allografts / Q000378:metabolism; D000920:Antibody-Dependent Cell Cytotoxicity; D000936:Antigen-Antibody Complex / Q000378:metabolism; D005260:Female; D005455:Fluorescent Antibody Technique / Q000379:methods*; D006084:Graft Rejection / Q000175:diagnosis* / Q000276:immunology; D006680:HLA Antigens / Q000378:metabolism; D006801:Humans; D007518:Isoantibodies / Q000378:metabolism; D007668:Kidney / Q000378:metabolism / Q000473:pathology; D016030:Kidney Transplantation; D008297:Male; D008875:Middle Aged
D000936:Antigen-Antibody Complex; D006680:HLA Antigens; D007518:Isoantibodies
Antibody-mediated rejection; donor-specific antibodies; immunocomplex capture fluorescence analysis