A method for the enrichment of live thyrotrophic pituitary cells is described. Pituitary glands of young male rats were removed into Earle's solution and dispersed in a 0·1% trypsin solution containing 0·5% bovine serum albumin, pH 7·2.
Nylon fibres (25 μm) were used for the separation of the thyrotrophic cells, by stringing them across a plastic frame which fitted a plastic Petri dish containing the cell suspension. The fibres were washed with light petroleum (b.p. 60–80 °C) and carbon tetrachloride, hydrolysed with 3 m-HCl for 30 min at room temperature and washed with distilled water and phosphate-buffered saline (pH 7·2). The fibres were treated with thyrotrophin releasing hormone (TRH) alone or in the presence of soluble carbodiimide solution. After incubation for 1 h at room temperature, the fibres were transferred to a new Earle's medium and cells were released from the fibres by plucking them with a needle. The separated thyrotrophic cells were identified by radioimmunoassay and by electron microscopy.
Using the above-mentioned methods, enrichment of thyrotrophic cells was obtained. Thus, the amounts of TSH, prolactin, LH and GH released, during 2 h of incubation, by 1·5 × 106 unseparated cells were 6·8 ± 0·65, 4·1 ± 0·47, 4·8 ± 0·52 and 5·2 ± 0·68 μg respectively, while the same number of purified thyrotrophic cells released 76·1 ± 0·42, 1·2 ± 0·3, 0·6 ± 0·35 and 1·6 ± 0·22 μg of the same hormones (means ± s.e.m.).
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Online ISSN: 1479-6805
Print ISSN: 0022-0795
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