ISOLATION, CHARACTERIZATION AND RADIOIMMUNOASSAY OF LUTEINIZING-HORMONE IN THE BRUSHTAIL POSSUM

Luteinizing hormone (LH) was purified from brushtail possum (Trichosur us vulpecula) pituitary glands. The purification procedure consisted o f ammonium sulfate precipitation followed by triazinyl-dye chromatogra phy, hydrophobic interaction chromatography and gel filtration. A yiel d of 10 mu g LH g(-1) pituitary with a recovery of 20% was obtained fr om 1400 pituitary glands (20.3 g). Contamination with possum follicle- stimulating hormone (FSH) was less than or equal to 0.05%. The amino a cid analysis and the N-terminal sequencing for 10 cycles revealed clos e homology with LH from other mammals. Minor amounts of LH that had be en truncated near the N-terminal were also detected. No contaminating proteins were found by amino acid sequencing. The potency of possum LH was 20% that of ovine LH in a receptor assay using possum testicular receptors and 4% that of ovine LH when bovine corpora lutea receptors were used. Possum LH was able to stimulate production of cyclic adenos ine 3',5'-monophosphate by bovine granulosa cells. A radioimmunoassay (RIA) for possum LH using I-125-possum LH and an antiserum raised agai nst ovine LH was developed. The RIA has a sensitivity of 0.15 ng mL(-1 ), a 50% displacement of 1.9 ng mL(-1) and a cross-reactivity of <0.02 % against possum FSH. Plasma concentrations were 0.24+/-0.04 ng mL(-1) (n = 8) and 0.39+/-0.12 ng mL(-1) (n = 8) in female and male possums respectively. Administration of mammalian gonadotrophin-releasing horm one (GnRH) and chicken GnRH II stimulated increases in plasma LH conce ntrations in male and female possums. When comparing LH responses with administration of mammalian GnRH or chicken GnRH II, plasma LH concen trations remained elevated for a longer period of time in males than i n females (P < 0.01); plasma LH concentrations also remained elevated for longer after mammalian GnRH than after chicken GnRH II (P < 0.01). Gonadectomy stimulated an increase in plasma concentrations of LH in both male (P < 0.01) and female (P < 0.05) possums. The rate of increa se in plasma LH concentrations in males was faster than that in female s. In summary, we have purified, partially characterized, and develope d a RIA for possum LH.