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Characterization and refinement of monoclonal anti‐human globulins that lack reactivity with human IgG4
Author(s) -
Howie Heather L.,
Collins Baodan,
Wang Xiaohong,
Kapp Linda,
Delaney Meghan,
Er Lay See,
Lebedev Jen.,
Zimring James C.
Publication year - 2020
Publication title -
transfusion
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.045
H-Index - 132
eISSN - 1537-2995
pISSN - 0041-1132
DOI - 10.1111/trf.15796
Subject(s) - epitope , monoclonal antibody , subclass , antibody , flow cytometry , monoclonal , arginine , epitope mapping , microbiology and biotechnology , chemistry , immunology , biology , amino acid , biochemistry
BACKGROUND Anti–red blood cell (RBC) alloantibodies consisting of only the immunoglobulin G (IgG) 4 subtype are typically considered clinically insignificant. A US Food and Drug Administration–approved monoclonal anti‐human globulin (16H8) is nonreactive with IgG4, which has been considered a benefit to avoid testing interference from IgG4. However, 16H8 also does not recognize two natural IgG3 variants (IgG3‐03 and IgG3‐13). Thus, 16H8 may miss clinically significant alloantibodies in some settings. STUDY DESIGN AND METHODS Novel mouse anti‐human IgG hybridomas were generated and screened for reactivity with 32 human variants of anti‐KEL1 across different IgG subtypes, as well as mutants to allow epitope mapping. Anti‐IgG reactivity was determined using KEL1+ RBCs bound by each IgG variant as targets. Binding of anti‐IgG was determined by flow cytometry. RESULTS 16H8 recognized an epitope involving amino acid 419, which is glutamate in IgG4, IgG3‐03, and IgG3‐13, explaining the lack of 16H8 reactivity with these subtypes/isoallotypes. A new monoclonal antibody (PUMA8) was isolated that, like 16H8, was nonreactive with IgG4 but without blind spots for known variants of IgG1, IgG2, or IgG3. PUMA8 recognized an epitope containing arginine at position 355, which is glutamine in IgG4. However, a recently described new IgG4 variant with an arginine at position 355 results in PUMA8 reactivity. CONCLUSION PUMA8 represents an alternative to 16H8 that avoids IgG4 but without blind spots for IgG3 variants. However, PUMA8 reacts with one recently described IgG4 variant. In addition to relevance to immunohematology, these studies highlight the importance of patient variation with regards to assay performance in an era of personalized medicine.