Abstract
Antipneumococcus plasma containing 0.4% trikresol is dialyzed in running tap water for 4 days. This will remove sufficient salts for the method to be described. The plasma and precipitate consisting mostly of fibrinogen, euglobulin and antibodies are removed from the dialyzing bags, sufficient sodium chloride or sodium sulphate is added to make the dialysate one-twentieth normal (50 cc. normal sodium chloride or sulphate to each liter) and placed in a cold room over night to lower the temperature to about 5°C. Twotenths percent trikresol is then added and the mixture adjusted to pH 5 to 5.1, using gamma dinitrophenol 0.025% solution as indicator. The antibodies are soluble at this pH and salt content. This mixture is allowed to remain at about 8°C. for 4 hours, during this time the insoluble substance fibrin, fibrinogen, euglobulin and the chill producing substance will have flocculated and settled to about 1/20 of the fluid volume. At this cold temperature it is then filtered clear through paper pulp (filtration is very rapid, 20 liters within a half hour). The filtrate is adjusted to pH 6.8 using para nitrophenol 0.1% solution as indicator, and allowed to stand over night at cool temperature for further precipitation, principally fibrin like substance at this pH. It is then filtered through paper pulp and trial tubes set up to determine the amount of distilled water necessary to precipitate the antibodies. This rarely requires more than two and a half volumes of distilled water. The required water is added and the precipitate allowed to settle in cool room over night. The following morning the supernatant fluid is decanted and the remainder of the precipitate centrifuged. The centrifuged precipitate is roughly estimated in cubic centimeters and a like amount of one percent sodium chloride containing 0.8% phenol is added and the precipitate dissolved.
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