| Enzyme-linked immunosorbent assay for rat hepatic triglyceride lipase. | |
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MedLine Citation:
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PMID: 3989393 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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A noncompetitive enzyme-linked immunosorbent assay to measure rat hepatic triglyceride lipase (H-TGL) was developed. Antibodies to rat H-TGL were purified from goat antisera by immunoadsorption on an H-TGL-Sepharose 4B column. Routinely, Immulon 2 Removawell strips were coated with the purified antibody overnight at 4 degrees C. After blocking the wells with bovine serum albumin (BSA) for 2 hr at room temperature, standards (0.85 ng/ml-13.1 ng/ml) or samples were added to the wells and were incubated with the bound anti-rat H-TGL overnight at 4 degrees C. The standards and samples had been pretreated with 5-20 mM SDS for 30 min at room temperature and were then diluted so that the final SDS concentration in the assay was 1 mM or less. The pretreatment with SDS was necessary to achieve maximal immunoreactivity. The sample incubation was followed by an overnight incubation at 4 degrees C with an anti-rat H-TGL-horseradish peroxidase conjugate. Rat H-TGL was detected by the color development after the addition of 0.4 mg/ml of o-phenylenediamine in 0.01% H2O2, 0.1 M citrate phosphate, pH 5.0. A linear relationship was obtained between absorbance at 490 nm and the amount of highly purified rat H-TGL used as a standard. Inclusion of 1 M NaCl in the assay buffer (1% BSA, 0.05% Tween 20, 10 mM phosphate, pH 7.4) during the sample and conjugate incubations minimized non-specific interactions. Recoveries of purified rat H-TGL added to a rat liver perfusate sample ranged from 98.6% to 103%.(ABSTRACT TRUNCATED AT 250 WORDS) |
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Authors:
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L A Cisar; A Bensadoun |
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Publication Detail:
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Type: Journal Article; Research Support, U.S. Gov't, P.H.S. |
Journal Detail:
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Title: Journal of lipid research Volume: 26 ISSN: 0022-2275 ISO Abbreviation: J. Lipid Res. Publication Date: 1985 Mar |
Date Detail:
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Created Date: 1985-06-13 Completed Date: 1985-06-13 Revised Date: 2007-11-14 |
Medline Journal Info:
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Nlm Unique ID: 0376606 Medline TA: J Lipid Res Country: UNITED STATES |
Other Details:
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Languages: eng Pagination: 380-6 Citation Subset: IM |
Export Citation:
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APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
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Animals Antibody Specificity Blood Proteins / analysis Electrophoresis, Polyacrylamide Gel Enzyme-Linked Immunosorbent Assay Lipase / isolation & purification* Liver / enzymology* Male Rats Rats, Inbred Strains |
| Grant Support | |
ID/Acronym/Agency:
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HL-14990/HL/NHLBI NIH HHS; HL-24873/HL/NHLBI NIH HHS |
| Chemical | |
Reg. No./Substance:
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0/Blood Proteins; EC 3.1.1.3/Lipase |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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