Ontogeny of corticotrophin-releasing factor (CRF) in the ovine fetal hypothalamus: use of multiple CRF antibodies.
In previous studies, using one particular antibody, immunohistochemical localization of corticotrophin-releasing factor (CRF) in ovine fetal brain was not possible before 90 days of gestation (term is approximately 150 days), although radioimmunoassay of hypothalamic extracts, using a different antibody, had shown CRF to be present from 63 days. The purpose of this study was to use a variety of CRF antibodies in both immunohistochemistry and radioimmunoassay to determine the presence and concentration of the CRF peptide as early in gestation as possible, and to determine whether more than one molecular size of CRF is detectable at any time in gestation. Seven different antibodies were used on hypothalamic tissue or extracts from seven adult sheep and 37 fetuses from 48 to 140 days of gestation. With one ovine CRF antibody (provided by Dr W. Vale, Salk Institute) immunohistochemical detection of CRF-labelled neurones and nerve fibres of the paraventricular nucleus and median eminence was possible from 49 days. The antibody with the greatest sensitivity in radioimmunoassay was one raised against human CRF, Ab-code R1 (provided by Dr E. Hillhouse, University of Newcastle upon Tyne). The hypothalamic contents of CRF (ng/whole hypothalamus) were 0.28 +/- 0.06 (mean +/- S.E.M.) (n = 4), 9.0 +/- 0.6 (n = 5), 14.3 +/- 0.6 (n = 5), 30.0 +/- 3.4 (n = 4) in fetuses at 48-50, 100-109 and 139-140 days of gestation and in adult sheep respectively. At all ages only one peak of CRF-like activity, which co-eluted with synthetic ovine CRF, was observed after chromatography of hypothalamic extracts, and assays performed with three different antibodies.(ABSTRACT TRUNCATED AT 250 WORDS)
Watabe
T
T
Howard Florey Institute of Experimental Physiology and Medicine, University of Melbourne, Parkville, Victoria, Australia.
Levidiotis
M L
ML
Oldfield
B
B
Wintour
E M
EM
D000818:Animals; D000906:Antibodies; D002850:Chromatography, Gel; D003346:Corticotropin-Releasing Hormone / Q000032:analysis / Q000276:immunology / Q000502:physiology*; D005865:Gestational Age; D007031:Hypothalamus / Q000737:chemistry / Q000196:embryology*; D007150:Immunohistochemistry; D011863:Radioimmunoassay / Q000379:methods; D012680:Sensitivity and Specificity; D012756:Sheep / Q000196:embryology*
D000906:Antibodies; D003346:Corticotropin-Releasing Hormone