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At least 109 records · Page 6Linked to original sources

Competitive binding assay for estrogen receptor in monolayer culture: measure of receptor activation potency.

MCF-7 cells were incubated with [3H]estradiol, unlabeled estradiol and various estrogens or antiestrogens to measure their relative binding affinity (whole-cell assay). Comparison of the values with those previously established on uterine cytosol with a DCC assay revealed a good parallelism for both steroid and diphenolic diethylstilbestrol based estrogens. On the contrary, in the whole-cell assay, antiestrogens and weak estrogens of the triphenyl- and gem-diphenylethylene categories always displayed low values which were in the order of magnitude found with weak steroid estrogens. This property was not due to a reduction of binding capacity, nor to the presence in some compounds of an ethoxyaminoalkyl side-chain (source of antiestrogenicity). In view of all these data, it was concluded that the present test would provide an estimate of the ability of a given compound to transform the receptor in a form which interacts with genomic sites involved in the regulation of "estrogenic-induced products" ("activation").

Binding, Competitive↗

A sensitive competitive binding assay for exogenous and endogenous heparins.

A new type of assay for heparins has been devised, in which the test material competes with 125I-labelled heparin for binding to protamine-Sepharose. The assay is very sensitive and will measure heparin concentrations down to 10 ng ml-1. It responds to both the degree of sulphation and the molecular weight of acidic polysaccharides, but is independent of their biological activities. It can be used to quantitate heparins in biological fluids after pretreatment of the samples with protease. In this way endogenous heparins were measured in normal human serum, plasma and urine. The assay is extremely versatile and has great potential for the investigation of endogenous and exogenous heparins.

Binding, Competitive↗

Competitive binding with putative Bo5 (CD5) cluster of monoclonal antibodies.

The relationship of seven monoclonal antibodies, putatively to the Bo5 (CD5) antigen, was tested. Five of the mAbs were confirmed to be directed against the Bo5 antigen. Three mAbs, CC29, BLT-1 and 8C11, effectively blocked binding to bovine PBM of mAb CC17, previously reported to be directed against this antigen. MAb 8-3F4 also blocked binding of mAb CC17, but less effectively than the others. MAbs IL-A67 and 79-5 did not inhibit binding of mAb CC17 because of antibody allelic specificity or technical reasons.

Animals↗

Competitive binding studies with multiple sites. Effects arising from depletion of the free radioligand.

Apparently anomalous behaviour arises if a radiolabelled drug and a non-radioactive drug compete for binding to a membrane-bound receptor when (a) there is severe depletion of the radiolabelled drug and (b) the non-radioactive drug binds to a heterogeneous population of binding sites. In extreme cases, binding of the non-radioactive drug to the sites of high affinity can be obscured completely. This phenomenon is illustrated by the binding of carbachol to muscarinic receptors, estimated by inhibition of a radiolabelled antagonist.

Animals↗

The anomalous behaviour of exogenous 25-hydroxyvitamin D in competitive binding assays.

The Vitamin D International External Quality Assessment Scheme (DEQAS) was established in 1989 to monitor the performance of assays for 25-hydroxyvitamin D (25-OHD) and 1,25-dihydroxyvitamin D (I,25(OH)(2)D). This is achieved through the quarterly distribution of five samples of human serum. Results are used to calculate an All-Laboratory Trimmed Mean and a Method Mean for each of the methods used by participants. In July 2005, participants were asked to assay serum to which 50.9 nmol of either 25-OHD(3) or 25-OHD(2) had been added as ethanolic solutions. The final concentration of ethanol in the serum was 0.7%. The distribution also included a sample of the original serum (OS) containing 0.7% pure ethanol. The percentage recoveries of exogenous 25-OHD(3) (R1) and 25-OHD(2) (R2) were calculated for each method. Results (OS nM, R1 and R2) were as follows: DiaSorin RIA (n=53); 39.2, 82.1%, 83.3%, DiaSorin Liason (n=16); 36.8, 81.4%, 88.6%, IDS RIA (n=21); 36.4, 54.2%, 29.1%, IDS OCTEIA (n=16); 47.3, 78.8%, 56.4%, Nichols Advantage (n=21); 58.9, 46.4%, 43.2%, HPLC (n=9); 42.6, 112.2%, 97.1%, LC-MS (n=4); 34.0, 111.5%, 118.1%. The IDS RIA and Nichols assays gave unexpectedly low recoveries. This does not appear to be a calibration problem or the effect of ethanol.

Binding, Competitive↗