Use of immunoaffinity chromatography for purification of 125I-labeled human prolactin
We assessed a simple method for purifying 125I-labeled human prolactin, taking advantage of the abundant supplies of monoclonal antibodies available. 125I-Labeled human prolactin purified by immunoaffinity chromatography is compared with that purified by gel filtration on Sephadex G-100. We used monoclonal antibodies to prolactin to prepare the affinity chromatography columns. Prolactin was radiolabeled by the Chloramine T method, purified by each of the above procedures, and the binding and displacement characteristics were studied in radioimmunoassays in which either monoclonal antibodies or a rabbit anti-prolactin serum was the first antibody. A nonimmune fraction of 125I-labeled prolactin that co-eluted with the immunoreactive hormone from Sephadex G-100 was removed by affinity chromatography, which increased the antibody binding of 125I-labeled prolactin in the radioimmunoassay in the absence of unlabeled antigen (B/T0, in percent) twofold or more, increased the assay sensitivity, and increased the slope of antigen displacement measured by the 50% intercept. Several advantages make this the purification method of choice.
|Authors||Stuart, M. C.;Boscato, L. M.;Underwood, P. A. :|
|Publisher Name||CLINICAL CHEMISTRY|
|URL link to publisher's version||http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=6821925|