Antipeptide polyclonal antibodies specifically recognize each human thyroid hormone receptor isoform.
We characterized four antipeptide polyclonal antibodies (abs) able to specifically recognize each thyroid hormone receptor (TR) isoform. The abs immunoprecipitated both the in vitro synthesized receptor and the receptor expressed in E. coli and their specificity was confirmed by competition studies and immunohistochemistry. Ab activity measured by enzyme-linked immunosorbent assay decreased after preabsorption of each ab with the immunizing peptide or the specific receptor protein expressed in E. coli. No specific activity was detectable in enzyme-linked immunosorbent assay, no nuclear staining was observed after affinity column immunoabsorption, and the specific bands obtained in Western blot analysis disappeared after preabsorption with the specific TR isoform expressed in E. coli. By immunohistochemical studies we detected coordinate expression of each receptor isoform in most tissues examined. However, in heart and muscle, the beta-isoform is expressed at a very low level compared to the alpha-isoform in spite of the significant TR beta mRNA levels previously demonstrated by Northern blot analysis. We also demonstrated a different pattern of distribution of alpha- and beta-isoforms in rat testis. In this tissue the TR alpha is significantly expressed in spermatogonia nuclei, but in spermatids the beta-isoform is predominant, and only the TR beta is detectable in mature spermatozoa.
Falcone
M
M
Department of Medicine, University of Chicago, Illinois 60637.
Miyamoto
T
T
Fierro-Renoy
F
F
Macchia
E
E
DeGroot
L J
LJ
D000818:Animals; D000906:Antibodies / Q000032:analysis / Q000276:immunology*; D000918:Antibody Specificity; D015152:Blotting, Northern; D015153:Blotting, Western; D002846:Chromatography, Affinity; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli; D006801:Humans; D007150:Immunohistochemistry; D007536:Isomerism; D008297:Male; D009132:Muscles / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D009206:Myocardium / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D011233:Precipitin Tests; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D011988:Receptors, Thyroid Hormone / Q000032:analysis / Q000235:genetics / Q000276:immunology*; D013737:Testis / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure
D000906:Antibodies; D012333:RNA, Messenger; D011988:Receptors, Thyroid Hormone