Abstract
The method is based upon a simple procedure for concentrating the anterior pituitary hormone 1 in the urine and injecting 2 or 3 small doses subcutaneously into immature female rats, 18-21 days old. The animals, including untreated normal controls, are autopsied on the second or third day at the latest and the diagnosis made from the gross changes in the reproductive organs. Serial sections are prepared from the ovaries, tubes, and uterus for confirmation of the macroscopic findings. The procedure shortens the time required for diagnosis to 36-48 hours instead of the usual 4-5 or more days.
Eight ounces of morning urine, preferably not over 8 hours old, are used in the test. Older specimens have been found satisfactory, however, and the results not vitiated by the addition of a preservative or by the reaction of the urine. A preservative such as ether tricresol (4 drops to each 100 cc. of urine) may be necessary in forwarding specimens from a distance.
Two and one-half volumes of 95% alcohol are added to the urine and the mixture placed in the ice chest at a temperature of 2-4°C. for several hours or over night to allow the precipitate to settle out. This step may be hastened by centrifuging the mixture as soon as the precipitate has separated. The precipitate is suspended in 3-4 cc. of physiological salt solution, shaken thoroughly with an equal volume of ether, centrifuged to remove the ether and the extraction repeated 2 or 3 times. The saline solution after centrifuging and containing the specific hormone freed from the estrus-producing or ovarian hormones is now ready for injection into rats.
Interpretation of the test is based upon the positive findings of enlarged ovaries with visible corpora hemorrhagica or protruding follicles and “blood points”.
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