Abstract
The following method for the extraction of gonadotropic hormone from human blood is a modification and improvement of the acid alcohol method previously reported from this laboratory. 1
With the old method only a faint Reaction I was demonstrable in 40 cc. of blood taken on the 8th to the 12th day of the cycle. With the new method it has been possible to obtain both strong Reaction I and III with the same amount of blood at a comparable point in the cycle.
Method. (1) 40 cc. of vein blood is mixed with 30 gm. of anhydrous sodium sulphate, stirred to dryness and pulverized. (2) 200 cc. of 60% alcohol, acidified with 50% acetic acid to pH 5, is then added to the powder and shaken for one hour in a mechanical shaker. (3) The mixture is centrifuged and the supernatant alcohol decanted and kept. (4) A similar second extraction of the residual sludge is made. (5) The 2 alcohol extracts are each concentrated by evaporation under a fan at room temperature to a volume of 50 cc. and then centrifuged. (The concentration can be hastened by vacuum distillation, temperature not to exceed 23°C.). (6) The precipitate and scum which separate during the concentration are removed and shaken for 10 minutes with 10–20 cc. of dilute NaOH, the final pH of the mixture not to exceed 8.5 (thymol blue) (A). (7) The two 50 cc. alcohol extracts are united and allowed to evaporate at room temperature (fan) almost to dryness (B). (8) The residue (B) is added to the aqueous alkaline extract of the first precipitate (A), then shaken and the pH of the whole adjusted to 8–8.5. The extract is then shaken vigorously for 5 minutes and allowed to stand for 10 minutes at room temperature (C).
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