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Production of recombinant murine‐human chimeric IgM and IgG anti‐Js b for use in the clinical laboratory
Author(s) -
Huang T.J.,
Reid M.E.,
Halverson G.R.,
Yazdanbakhsh K.
Publication year - 2003
Publication title -
transfusion
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.045
H-Index - 132
eISSN - 1537-2995
pISSN - 0041-1132
DOI - 10.1046/j.1537-2995.2003.00393.x
Subject(s) - clone (java method) , antibody , microbiology and biotechnology , flow cytometry , biology , recombinant dna , complementary dna , chimera (genetics) , antigen , transfection , fusion protein , immunoglobulin m , virology , immunoglobulin g , immunology , cell culture , gene , genetics , biochemistry
BACKGROUND : Directly agglutinating MoAbs are more useful than IgG MoAbs of murine origin for typing RBCs from donors and patients. The molecular manipulation and conversion of a murine IgG MoAb into mouse‐ human chimeric IgM and IgG antibodies are described. STUDY DESIGN AND METHODS : cDNA encoding the variable heavy‐ and light‐chain genes of a murine hybridoma anti‐Js b cell line (MIMA‐8) were cloned into human IgM or IgG expression vectors, which were then separately stably transfected into SP2/0 ‐Ag14 B‐cells. The secreted antibodies were screened by ELISA and analyzed by flow cytometry and hemagglutination. RESULTS : Forty percent (16 of 40) of the stable clones secreted IgM and 66 percent (12 of 18) of the stable clones secreted IgG. The chimeric IgM from the highest expressing clone reacted 4+ in LISS at room temperature. The chimeric IgG from one clone reacted 4+ by the IAT, resembling the specificity of the original murine antibody. Both manipulated MoAbs reacted specifically with RBCs as assessed by flow cytometry. CONCLUSION : Human‐mouse chimeric IgM and IgG from a murine IgG MoAb anti‐Js b has been successfully engineered for use in the clinical laboratory. This approach can potentially be used to manipulate other murine MoAbs to blood group antigens into more clinically useful human isotypes.