Purification of rat prolactin : Development of an homologous radioimmunological assay and comparison with the NIAMDD system

Abstract

the pituitaries dissected. Using a thorough fractionation procedure, the other pituitary hormones were eliminated. To obtain a PRL preparation with high biological activity, gel filtration on Sephadex G 100 was used. The prepared PRL was studied using physicochemical, biological and immunological methods. From gel filtration data the molecular weight of the PRL thus prepared was approximately 26 000. Using a radioreceptor assay the activity of the preparation was two times higher than the NIAMDD preparation of PRL (NIAMDD-RP,). Immunofluorescence studies revealed that rabbit antiserum raised against this PRL had affinity for the PRL cells in ovine, bovine and rat pituitaries but none for the other types of pituitary cells. A double antibody radioimmunoassay was developed using the above hormone and its antiserum. Standard curves were plotted over the range of 16 to 400 pg. Rat FSH, LH, TSH and GH did not react in this assay nor did plasma proteins. The intra-assay coefficient of variation was 10 p. 100 and the between assay 16 p. 100. The rat prolactin radioimmunoassay kit distributed by the National Institute of Arthritis and Metabolic Diseases (NIAMD) was also examined. Prolactin levels in rat serum measured by the NIAMD-radioimmunoassay were twice those measured by our system. The apparent differences in serum levels of PRL simply reflected the lower immunologic activity of the NIAMD prolactin.