THE UNIVERSITY OF MANITOBA STUDIES ON OVINE PLACINTAL ...

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All ho¡¡none preParationFr pituitary and placental
extracts, Êerum, and oPL ttracer weie diluted in 0'.01-M phosPhate-
buffered saline (PBS) | pH 7.6r containing 2.52 BSA, and a 0'1 ml
sample ( or 0.2 ml sanple) v¡as taken for assay. Approximately
30,000 to 35,000 cpÍr ovine [125r] iodo-PI. in 0.1 ml buffer,
0.1 ml diluted antiserun to oPL (1,,/501000 dilution) r 0'1 mI
oPL standard or assay sampleso and 0.5m1 PBS t pH 7.4, containing
2.58 of BSA were added to glass tubes (L2 x 75 cm). After a
¿i ¡'
72-in incubat.ion at 4 C, 0.1 ¡n1 sheep anti-rabbit galnrnaglobulin
serun (1,/35 dilution) and 0'I ml ndrmal rabbit serum
lI/350 dilution) were added. After a further 24-h incubation
perio¿I in the cold, the precipitates formed hrere centrifuged
at 780 X g for 30 min, and the supernatant was decanted'
The radioactivity of the precÍpitates was then counted in a
LKB ganuna-counter (modej- 8000).
Pituitarv and Placental Extracts
Pituitary and placental- tissues from various species
were homogenized with 0.1 M NH4HCO3, pH 8"4 at 4C \'ríth a
tissue to buffer ratio of t:5 (wt,/voI), stirred overnight'
and centrifuged at l-5,000 x I for 20 min; the Pellet was
discarded and the supernate was stored at -20 C until assay'