A sensitive sandwich enzyme-linked immunosorbent assay of rat apolipoprotein A-I: effect of various sample treatments on apolipoprotein A-I immunoreactivity and an application to young and aged rat sera.
A highly sensitive sandwich enzyme-linked immunosorbent assay for rat apo A-I was developed. Samples and standards were added to each well of microtiter plates precoated with immunoaffinity-purified IgG. Bound apo A-I was detected with immunoaffinity-purified Fab'-horseradish peroxidase conjugate by a colorimetric method. The sensitivity reached 2.5 pg/well, and the working range for the measurement of serum apo A-I concentration was 0.1 to 1.0 ng/well. The mean intra- and interassay coefficients of variation were 2.8 and 4.1%, respectively. The epitopes of apo A-I in serum were effectively exposed by the use of 6 mol/liter guanidine.HCl. Serum apo A-I concentrations in 36- to 40-week-old rats (62.3 +/- 8.6 mg/dl, mean +/- SD, n = 16) were significantly higher (P less than 0.05) than those in 8- to 12-week-old rats (55.1 +/- 4.3 mg/dl, n = 9). But the age-related change of serum apo A-I was much smaller than that of serum apo E. Apo A-I was contained in smaller HDL particles (or HDL2) in normal rat serum.
Chiba
H
H
Department of Laboratory Medicine, Hokkaido University School of Medicine, Sapporo, Japan.
Mitamura
T
T
Matsuno
K
K
Kobayashi
K
K
D000375:Aging / Q000097:blood; D000818:Animals; D016632:Apolipoprotein A-I / Q000032:analysis / Q000378:metabolism*; D001057:Apolipoproteins E / Q000097:blood; D002784:Cholesterol / Q000097:blood; D003902:Detergents; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; D007202:Indicators and Reagents; D008297:Male; D011136:Polysorbates; D051381:Rats; D011919:Rats, Inbred Strains
D016632:Apolipoprotein A-I; D001057:Apolipoproteins E; D003902:Detergents; D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; D007202:Indicators and Reagents; D011136:Polysorbates; D002784:Cholesterol