A sensitive immunoassay for rat fatty acid translocase (CD36) using phage antibodies selected on cell transfectants: abundant presence of fatty acid translocase/CD36 in cardiac and red skeletal muscle and up-regulation in diabetes
| Autor: | M M, Pelsers, J T, Lutgerink, F A, Nieuwenhoven, N N, Tandon, G J, van der Vusse, J W, Arends, H R, Hoogenboom, J F, Glatz |
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| Rok vydání: | 1999 |
| Předmět: |
CD36 Antigens
Male Membrane Glycoproteins Myocardium Immunoglobulin Variable Region Organic Anion Transporters Enzyme-Linked Immunosorbent Assay Antibodies Viral Transfection DNA Fingerprinting Cell Line Rats Antibody Specificity Peptide Library Diabetes Mellitus Animals Humans Bacteriophages Rats Wistar Muscle Skeletal Immunoglobulin Fragments Research Article |
| Zdroj: | The Biochemical journal. 337 |
| ISSN: | 0264-6021 |
| Popis: | The rat membrane protein fatty acid translocase (FAT), which shows sequence similarity to human CD36 (a membrane protein supposedly involved in a variety of membrane processes), is implicated in the transport of long-chain fatty acids across cellular membranes. To set up an immunoassay for quantification of FAT in different tissues, we isolated a series of anti-FAT antibodies by panning a large naive phage antibody library on FAT-transfected H9c2 cells. All seven different phage antibody fragments isolated reacted specifically with FAT, and most likely recognize the same or closely located immunodominant sites on FAT, as a competitive monoclonal antibody (mAb) (CLB-IV7) completely blocked the binding of all these phage antibodies to cells. A sandwich ELISA was set up using mAb 131. 4 (directed against purified CD36 from human platelets) as capture antibody and phage antibodies and anti-phage sera as detector. With this ELISA (sensitivity 0.05 microgram/ml), the FAT content in isolated cardiomyocytes was found to be comparable with that of total heart ( approximately 3 mg/g of protein), while liver tissue and endothelial cells were below the detection limit ( |
| Databáze: | OpenAIRE |
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