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A murine monoclonal antibody against Kx protein which reacts also with β‐spectrin
Author(s) -
Carbonnet,
Blanchard,
Hattab,
Cochet,
Petit-Leroux,
Loirat,
Cartron,
Bertrand
Publication year - 2000
Publication title -
transfusion medicine
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.471
H-Index - 59
eISSN - 1365-3148
pISSN - 0958-7578
DOI - 10.1046/j.1365-3148.2000.00245.x
Subject(s) - spectrin , monoclonal antibody , glycoprotein , epitope , red blood cell , antibody , antigen , microbiology and biotechnology , membrane , flow cytometry , chemistry , band 3 , red cell , membrane protein , biochemistry , biology , immunology , cell , medicine , cytoskeleton
Kx is a polytopic membrane protein of human erythrocytes carrying the Kx blood group antigen, which is deficient in rare patients with McLeod syndrome. Kx is disulphide bond linked to the Kell glycoprotein, which is a bitopic type II membrane protein carrying the Kell blood group antigen. Mice immunized with a synthetic peptide predicted to be located on the second external loop of Kx produced a monoclonal antibody called 3E12 which does not recognize red cells with common Kell phenotype by agglutination and flow cytometry. 3E12 recognizes the Kx protein and the spectrin β‐chain on western blots, the affinity for these two proteins being lowered with increasing ionic strength. Linear epitopes recognized by 3E12 are E 116 EIEKE 121 and L 484 AQELEKE 491 on the Kx protein and spectrin β‐chain, respectively. To quantify the relative amount of Kx in Empigen BB extracts of red cell membranes, an ELISA for Kx was set up which showed conclusively that (i) there is less Kx in membranes of K 0 individuals (lacking the Kell glycoprotein) than in membranes of common individuals, and (ii) that all common individuals, typed as K+k–, K–k+ and K+k+, have the same amount of Kx on their red cell membranes. When an erythrocyte membrane detergent extract from one K 0 individual was chromatographed on an immobilized 3E12 column, a minute amount of authentic Kell glycoprotein was recovered in acid eluted fractions, indicating that at least the K 0 individual under study may still produce some Kell protein.