Linked Life Data

1790172

Source:http://linkedlifedata.com/resource/pubmed/id/1790172

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
5
pubmed:dateCreated
1992-4-2
pubmed:abstractText
The determination of enzyme levels in cellular extracts by active site titrations or by catalytic activity measurements is relevant in both science and medicine. However, these techniques assume that enzymes exhibit the same response in crude sample matrices as they do in the purified state. We report here an example of how an enzyme-linked immunosorbent assay (ELISA) was used to determine the true enzyme concentration which was compared to the effective enzyme concentration obtained by ligand binding and catalytic assay methods in a crude bacterial cell extract. Rabbit antibodies specific for Lactobacillus casei thymidylate synthase (TS) were used to develop a highly specific and sensitive heterogeneous noncompetitive ELISA assay with a typical detection limit of 1.4 fmol of TS (100 pg) and a dynamic working range of 3 orders of magnitude. The antibodies showed identical responses for TS, its inhibitory binary complex with 5-fluoro-2'-deoxyuridylate, and its inhibitory ternary complex with 5-fluoro-2'-deoxyuridylate and 5,10-methylenetetrahydrofolate in the immunoassay. L. casei cell-free extracts were subjected to extraction with CM-Sephadex and the various fractions were analyzed by ELISA, active-site titrations, and catalytic assays which demonstrated that assays which assumed full catalytic or ligand-binding competence underestimated the true enzyme level.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:issn
1043-1802
pubmed:author
pubmed:issnType
Print
pubmed:volume
2
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
327-32
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed:articleTitle
Comparison of ELISA with activity and ligand-binding methods for the determination of thymidylate synthase concentration.
pubmed:affiliation
Department of Chemistry, University of South Carolina, Columbia 29208.
pubmed:publicationType
Journal Article, Comparative Study, Research Support, U.S. Gov't, P.H.S.