Mouse Prolactin Obtained by Pituitary Organ Culture in a Serum-Free Medium

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Mouse prolactin was purified by organ culture of pituitaries and electrophoresis of the medium. Mouse pituitaries were organ-cultured in 9-cm Petri dishes containing Waymouths medium (MB 752/1) supplemented with penicillin (50units/ml), streptomycin (100μg/ml), and bovine insulin (0.12 I.U./ml) for 8 days. Prolactin-rich culture medium was half-saturated with ammonium sulfate and centrifuged. The pellet was subjected to analytical disc electrophoresis (10% acrylamide). Gels were sectioned into 2-mm segments. Prolactin was eluted in 0.04M phosphate buffer (pH 7.2), dialyzed and lyophylized. Two hundred and forty ml medium in which 360 pituitaries were cultured yielded 29.3mg lyophylized mouse prolactin. Although the preparation contained 2 other bands on acrylamide gel disc electrophoresis, a single precipitin line was seen in immunodiffusion and immunoelectrophoresis, showing the identity of their antigenicity. From these results, two other proteins in the preparation were suggested to be deamidated prolactin.
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