Characterization of a monoclonal antibody (HB-22) and development of an ELISA for human apolipoprotein A-I.
We describe the production and characterization of a high-affinity monoclonal antibody, HB-22, for apolipoprotein (apo) A-I, a major protein of human high-density lipoproteins (HDL). Including Tween 20 in the reaction mixture increased the binding capacity of HB-22 to apo A-I. HB-22 showed monospecific reactivity with HDL or apo A-I, displaying no cross-reactivity with apo A-II, intermediate-, low-, or very-low-density lipoproteins. Immunoaffinity columns with HB-22 (in the absence of Tween 20) showed an immunosorbent capacity of 80 micrograms of apo A-I per milligram of antibody. The immunosorbent capacity of HB-22 for apo A-I was similar in plasma samples from normolipidemic, hypercholesterolemic, or hypertriglyceridemic patients. Comparative binding studies demonstrated that compared with other available monoclonal apo A-I antibodies, HB-22 had the greatest apparent affinity for binding to HDL. A competitive ELISA developed by utilizing HB-22 could detect as little as 20 ng of apo A-I in the reaction mixture. The intra- and interassay CVs of the ELISA were 5.4% and 9.5%, respectively.
Rothwell
T C
TC
Hycor Biomedical Inc., Garden Grove, CA 92641, USA.
Kamanna
V S
VS
Jin
F Y
FY
Koren
E
E
Foley
T
T
Kashyap
M L
ML
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000276:immunology; D000918:Antibody Specificity; D016632:Apolipoprotein A-I / Q000097:blood* / Q000276:immunology; D001667:Binding, Competitive; D002846:Chromatography, Affinity; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D006937:Hypercholesterolemia / Q000097:blood; D015228:Hypertriglyceridemia / Q000097:blood; D007163:Immunosorbent Techniques; D008075:Lipoproteins, HDL / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D010084:Oxidation-Reduction; D011136:Polysorbates
D000911:Antibodies, Monoclonal; D016632:Apolipoprotein A-I; D008075:Lipoproteins, HDL; D011136:Polysorbates