| Autor: |
A Barkia, C Martin, P Puchois, J C Gesquiere, C Cachera, A Tartar, J C Fruchart |
| Jazyk: |
angličtina |
| Rok vydání: |
1988 |
| Předmět: |
|
| Zdroj: |
Journal of Lipid Research, Vol 29, Iss 1, Pp 77-84 (1988) |
| Druh dokumentu: |
article |
| ISSN: |
0022-2275 |
| DOI: |
10.1016/S0022-2275(20)38560-6 |
| Popis: |
Apolipoprotein A-I (apoA-I), the major protein component of human high density lipoprotein, appears intracellularly as an intermediate precursor (proapoA-I) with a hexapeptide extension (Arg-His-Phe-Trp-Gln-Gln) at its amino terminus. To investigate the regulation of processes that regulate plasma apoA-I levels, a sensitive and simple assay for proapoA-I is required. We describe a specific enzyme-linked immunosorbent assay (ELISA) for quantification of proapoA-I using monospecific rabbit antibodies raised against the peptide: Arg-His-Phe-Trp-Gln-Gln-Asp-Glu-Pro. The monospecificity of antibodies to propeptide has been checked and no cross-reaction with mature apoA-I has been found although three first mature apoA-I amino acids (Asp-Glu-Pro) were included in the immunizing peptide. The assay is a non-competitive sandwich ELISA in which polystyrene microtiter plates were used with antibodies to propeptide adsorbed on the wells. After incubation with plasma samples, the bound proapoA-I was revealed by labeled rabbit polyclonal antibodies directed against mature apoA-I. The working range was 10 to 100 ng/ml, recovery of proapoA-I added to plasma was 94.6 to 106.5%, and the intra- and interassay coefficients of variation were 3.8% and 7.9%, respectively. A delipidation step using diisopropylether-n-butanol was necessary to expose antigen sites of proapoA-I in native lipoproteins. Mean level of proapoA-I in normal subjects was 87 +/- 15 micrograms/ml. It represented 7.1% of total apoA-I while in Tangier serum it represented 29%. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
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