A monoclonal antibody to the rat nuclear triiodothyronine receptor: production and characterization

Endocrinology
M LuoJ H Dussault

Abstract

The nuclear T3 receptor (NTR) was affinity-labeled with bromoacetyl-[125I]T3, purified by preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and used to immunize BALB/c mice. Spleen cells from one strongly immunoreactive mouse were fused with Sp2 mouse myeloma cells, and 328 hybridomas were screened by a dot-blot immunoassay using as antigen, a preparation of NTR partially purified by diethylaminoethyl-Sephadex chromatography. Four positive cultures were thus found; three of which were confirmed by comparing Western blotting patterns with the electrophoretic mobility of the affinity-labeled NTR. One of these 3 hybridomas was further subcloned by limiting dilution and gave rise to the 2B3 clone, which produces an immunoglobulin of the immunoglobulin G1 subclass. Several lines of evidence indicated that the 2B3 monoclonal antibody was indeed directed against the NTR. The antibody recognized a protein with the same electrophoretic mobility as the affinity-labeled receptor. Thus, Western blotting revealed a predominant protein with a mol wt of 57,000 and a less abundant 45,000 component on sodium dodecyl sulfate gels, and multiple isoelectric variants of the 57,000 protein, with a predominant form at pI 6.2, wer...Continue Reading

Citations

Mar 1, 1989·Brain Research. Developmental Brain Research·M LuoJ H Dussault
Feb 3, 1992·Neuroscience Letters·J PuymiratJ H Dussault
Jan 1, 1991·Thyroid : Official Journal of the American Thyroid Association·J PuymiratJ H Dussault
Nov 15, 1989·Biochemical and Biophysical Research Communications·E D SchmidtW H Lamers
Aug 24, 1999·Journal of Neurobiology·I Barakat-Walter
Dec 1, 1989·Journal of Endocrinological Investigation·L J DeGrootE Macchia

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