A potential complication in the use of monoclonal antibodies: inhibition of apoB-mediated receptor binding by an anti-apoE antibody.
Monoclonal antibody (Mab) 1D7 is specific for human apolipoprotein (apo) E and blocks binding of lipid-associated apoE to the low density lipoprotein (LDL) receptor. We report here that 1D7 can also block the binding of apoE-free LDL to the LDL receptor. The inhibition of LDL-receptor binding is not due to immunological cross-reactivity between the anti-apoE Mab and apoB, the ligand responsible for the interaction of LDL with the LDL receptor: 1) Mab 1D7 did not react with apoE-depleted LDL; 2) the LDL receptor binding inhibitory activity of 1D7 immunoglobulin G (IgG) preparations could be dissociated from the anti-apoE activity; 3) the inhibition was maintained when the fibroblasts were preincubated with the 1D7 IgG, extensively washed, and only then exposed to 125I-labeled LDL. Rather, it appears that 1D7 recognizes mouse apoE, that mouse apoE-1D7 immune complexes contaminate 1D7 IgG preparations and that the contaminating mouse apoE can compete with 125I-labeled LDL for the LDL receptor. We have demonstrated mouse apoE in IgG preparations of 1D7 but not in those of other anti-apoE Mabs that do not influence LDL-receptor binding. Precipitation of 1D7 IgG with NH4SO4 eliminates both apoE and the capacity of 1D7 to block LDL receptor binding. Finally, mouse apoE can be isolated by immunoaffinity chromatography of mouse serum on immobilized 1D7 Mab. As this is probably not a unique case, the observation has important implications for the use of Mabs as structural probes.
Maurice
R
R
Laboratoire du Métabolisme des Lipoprotéines, Institut de Recherches Cliniques de Montréal, Québec, Canada.
Marcel
Y L
YL
Innerarity
T L
TL
Milne
R W
RW
D000911:Antibodies, Monoclonal; D001053:Apolipoproteins / Q000276:immunology / Q000378:metabolism; D002478:Cells, Cultured; D002846:Chromatography, Affinity; D003429:Cross Reactions; D004591:Electrophoresis, Polyacrylamide Gel; D005347:Fibroblasts / Q000166:cytology; D006801:Humans; D008077:Lipoproteins, LDL / Q000378:metabolism*; D011973:Receptors, LDL / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D001053:Apolipoproteins; D008077:Lipoproteins, LDL; D011973:Receptors, LDL