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Next Generation Sequencing B Cell Receptor Reveals B Cell Clones Encoding HLA Antibody in Transplant Recipients

X. Zhang

HLA and Immunogenetcis Laboratory /CTC, Cedars Sinai Medical Center, Los Angeles, CA

Meeting: 2020 American Transplant Congress

Abstract number: 546

Keywords: Antibodies, B cells, HLA antibodies

Session Information

Session Name: B-cells / Antibodies /Autoimmunity

Session Type: Oral Abstract Session

Date: Saturday, May 30, 2020

Session Time: 3:15pm-4:45pm

 Presentation Time: 4:27pm-4:39pm

Location: Virtual

*Purpose: The development of HLA donor specific antibodies (DSA) increases the risk of antibody mediated rejection and is an independent predictor of poor patient survival. The antibody specificity is defined by the single antigen beads-based assay. However, this assay cannot detect antibodies against unknown antigens, neither can it detect memory B cells. Antibodies are encoded by the B cell receptor genes (BCR) and antibody diversity is achieved by vast clone types of BCR repertories. Recent advances in next generation sequencing allows characterizing vast clone types and expression levels of BCR repertoire. The aim of this study is to characterize the BCR clone types in transplant recipients and determine if development of de novo DSA can be detected by changes of BCR clone types.

*Methods: We sequenced BCR in recipients who developed de novo DSA post-transplant. For bulk BCR heavy chain sequencing, mRNA was isolated from peripheral blood mononuclear cells before transplant and after development of de novo DSA. The BCR heavy chain gene was amplified by PCR and sequenced using miSeq. To obtain the paired information of the BCR heavy chain and light chain, B cells were enriched and BCR mRNA was single cell sequenced by using 10X Genomics. To generate antibodies, paired heavy chain and light chain genes were expressed in 293 cells and the specificity of expressed antibodies was determined by the single antigen beads-based assay.

*Results: Changes of BCR clone types were detected after recipients developed de novo DSA via analysis of the heavy chain V-D-J usage. The heavy chain CDR3, the most polymorphic region of the BCR genes, was enriched with certain unique CDR3 sequences in samples with de novo DSA. Expression of the BCR heavy chain gene encoding the most abundant CDR3 sequence together with the paired light chain generated an antibody against an HLA antigen. Of note, the specificity of the expressed antibody is different from the one detected in the recipient ‘s serum, suggesting that the BCR sequencing can detect B cells clones before the antibody can be detected in the serum.

*Conclusions: Development of de novo DSA is associated with changes of B cell clone types. NGS sequencing BCR may be used to detect memory response before the antibody can be detected in peripheral blood.

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To cite this abstract in AMA style:

Zhang X. Next Generation Sequencing B Cell Receptor Reveals B Cell Clones Encoding HLA Antibody in Transplant Recipients [abstract]. Am J Transplant. 2020; 20 (suppl 3). https://atcmeetingabstracts.com/abstract/next-generation-sequencing-b-cell-receptor-reveals-b-cell-clones-encoding-hla-antibody-in-transplant-recipients/. Accessed May 10, 2025.

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