A particle concentration fluorescence immunoassay for Lp(a).
The quantitation of Lp(a) by immunoassay presents a major technical problem, because the molecular mass of the (a) protein of Lp(a) can vary between 419,000 and 838,000 Da (Gaubatz et al. (1990) J. Lipid Res. 31, 603-612), and this variability is determined by at least 24 alleles of the (a) gene. In an attempt to overcome this problem, we have developed an assay that is independent of variation of the size of (a). The assay utilizes a mixture of monoclonal antibodies to (a) which do not react to plasminogen or to apolipoprotein (apo) B. These antibodies are bound to inert microscopic beads to capture the Lp(a) particles. Subsequently, a fluorescein-labeled monoclonal antibody to apo B is used for detection and quantitation. The assay is done with special microtiter plates containing filters so that the particles can be thoroughly washed after capture on the microbeads. Because Lp(a) particles contain only one apo B particle and the molecular weight of apo B is constant, the assay is not affected by variation in the size of apo(a). By binding the mixture of monoclonal antibodies to inert beads, it is possible to greatly increase the amount of antibody bound to an exposed surface and thus increase the sensitivity of the assay. A mixture of monoclonal antibodies can be used to increase the affinity of the capture step of the assay. The assay can be completed in 4 h and has a wide working range.(ABSTRACT TRUNCATED AT 250 WORDS)
Kottke
B A
BA
Mayo Clinic, Rochester, MN 55905.
Bren
N D
ND
D000818:Animals; D000911:Antibodies, Monoclonal; D001055:Apolipoproteins B / Q000032:analysis / Q000276:immunology; D005069:Evaluation Studies as Topic; D005455:Fluorescent Antibody Technique; D006801:Humans; D017270:Lipoprotein(a) / Q000097:blood* / Q000235:genetics / Q000276:immunology; D051379:Mice; D008970:Molecular Weight; D010641:Phenotype
D000911:Antibodies, Monoclonal; D001055:Apolipoproteins B; D017270:Lipoprotein(a)