Luteinizing hormone (LH) was isolated from camel hypophyses by extraction, precipitation and successive ion exchange chromatography, and gel filtration. Gel electrophoresis showed three main bands at 27·5-29, 17-18 and 15-16 kDa. Biological activity of the purified product (camLH) was assayed by evaluating testosterone production of rat Leydig cells after stimulation with the isolated camLH and with porcineLH. Dose response curves indicated that the camLH and the reference porcineLH had the same potency. Immunoreactivity was tested in a heterologous radioimmunoassay (RIA) system using an anti-ovineLH serum. The camLH standard curve showed close parallelism with that of the ovineLH used as a reference. The two curves had median values (B/Bo = 50%) of 449·5 ± 17·4 and 822·0 ± 69·2 pg tube-1 (mean ± S.E.M.) with sensitivities (B/Bo = 90%) of 58·2 ± 7·0 and 44·8 + 8·8 pg tube-1. The purified product obtained after gel-filtration was injected into rabbits to produce the specific anti-camLH serum which was then applied to the development of the new RIA which uses the isolated camLH as a standard reference and iodinated ovineLH as labelled antigen. Sensitivity was 57·7 ± 6·4 pg tube-1 with a median value of 238·0 ± 15·5 pg tube-1. Specificity was tested using ovine follicle stimulating hormone (FSH) as interferant. The cross-reactivity was 100% for camLH and 0·3% for ovineFSH. The anti-camLH serum showed good sensitivity and specificity in the heterologous RIA and appeared suitable for the development of a more specific homologous RIA system.
ISOLATION OF LUTEINIZING-HORMONE, ANTISERUM PRODUCTION AND THE DEVELOPMENT OF A RADIOIMMUNOASSAY FOR CAMELS
GABAI, GIANFRANCO;BONO, GABRIELE
1994
Abstract
Luteinizing hormone (LH) was isolated from camel hypophyses by extraction, precipitation and successive ion exchange chromatography, and gel filtration. Gel electrophoresis showed three main bands at 27·5-29, 17-18 and 15-16 kDa. Biological activity of the purified product (camLH) was assayed by evaluating testosterone production of rat Leydig cells after stimulation with the isolated camLH and with porcineLH. Dose response curves indicated that the camLH and the reference porcineLH had the same potency. Immunoreactivity was tested in a heterologous radioimmunoassay (RIA) system using an anti-ovineLH serum. The camLH standard curve showed close parallelism with that of the ovineLH used as a reference. The two curves had median values (B/Bo = 50%) of 449·5 ± 17·4 and 822·0 ± 69·2 pg tube-1 (mean ± S.E.M.) with sensitivities (B/Bo = 90%) of 58·2 ± 7·0 and 44·8 + 8·8 pg tube-1. The purified product obtained after gel-filtration was injected into rabbits to produce the specific anti-camLH serum which was then applied to the development of the new RIA which uses the isolated camLH as a standard reference and iodinated ovineLH as labelled antigen. Sensitivity was 57·7 ± 6·4 pg tube-1 with a median value of 238·0 ± 15·5 pg tube-1. Specificity was tested using ovine follicle stimulating hormone (FSH) as interferant. The cross-reactivity was 100% for camLH and 0·3% for ovineFSH. The anti-camLH serum showed good sensitivity and specificity in the heterologous RIA and appeared suitable for the development of a more specific homologous RIA system.
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