DEFENSIVE FERMENTS 249 



Determination of Antiferment in Blood Serum. The determination of 

 antitrypsin by the Fuld-Gross method is satisfactory for this purpose. This 

 requires in addition to blood serum taken preferably in the morning before 

 the patient's breakfast, solutions of casein, acetic acid, and trypsin. The 

 solution of casein is made by dissolving i gram casein in 100 c.c. N/io NaOH 

 with the aid of slight heating; the solution is neutralized with N/io NaCl and 

 made up to 500 c.c. with 0.85 per cent. NaCl. The acetic acid solution is made 

 by mixing 5.0 c.c. acetic acid with 45.0 c.c. alcohol and 50.0 c.c. water. The 

 trypsin solution is made by dissolving 0.5 gram trypsin (Griibler) in 50.0 c.c. 

 0.85 per cent. NaCl and 0.5 c.c. normal soda solution; this is diluted ten times 

 with saline. The patient's serum must be fresh and should be diluted with 

 salt solution so as to make a 2 per cent, solution. 



The trypsin is titrated as follows: Place in a series of test tubes o.i, 0.2, 

 0.4, 0.6, 0.8, and i.o c.c. of the trypsin solution. Add 2.0 c.c. of casein solution 

 to each tube, shake and incubate for one-half hour at 50 C. Add three or 

 four drops of the acetic acid solution to each tube and note the precipitation 

 (cloudiness) which appears in the course of a few minutes. The tube which 

 remains perfectly clear contains enough trypsin to digest 2.0 c.c. of the casein 

 solution. For testing the antitryptic content of the serum add 0.5 c.c. of the 

 2 per cent, solution of serum to each of six small test tubes. Then add to 

 each tube in series, increasing amounts of the trypsin solution, beginning with 

 the largest dose that completely digested the casein, and increasing in each 

 tube by o.i c.c. Add 2.0 c.c. of casein solution to each tube and make up to 

 equal volumes with normal saline. Shake and incubate for one-half hour at 

 50 C.; then add three or four drops of the acetic acid solution to each tube 

 and observe as before. The amount of trypsin which is inhibited by the 

 serum is determined by the lack of complete digestion, as shown by the acetic 

 acid precipitation. A control series should be set up with the pooled sera of 

 normal individuals. Jobling and his associates made the test somewhat more 

 accurate by filtering after the incubation and then determining quantitatively 

 the non-coagulable protein nitrogen. 



The Abderhalden Test. In the discussion of the specificity of fer- 

 ments, we pointed out that Abderhalden had assumed that the entry of 

 cellular and other proteins in the circulation could lead to the formation, 

 or increase, of ferments which have as their specific character the prop- 

 erty of digesting the antigenic protein. The test is based fundamentally 

 upon a mixture of serum and antigenic substance and the determina- 

 tion of the formation of diffusible protein products. He regarded the 

 ferments as protective, inasmuch as they could break down and aid 

 in the elimination o>f substances essentially foreign in nature. The 

 technic of the test has been carefully reviewed by Bronfenbrenner in 

 Vol. I of the Journal of Laboratory and Clinical Medicine, and we call 

 particular attention to certain modifications that have been offered by 

 Retinger in Volume XXII of the Archives of Internal Medicine. The 

 following brief description of the test is given in order to provide an 

 outline of the general principles. 



The materials essential for the test are the serum or plasma of the 

 patient, the substratum, dialyzing tubes and flasks, carefully distilled water, 

 clean test tubes and ninhydrin. The blood is withdrawn before the patient's 

 Ibreakfast in order to obtain blood at a time when no dialyzable products of 

 intestinal digestion are present. It may be taken into paraffin-coated centri- 

 fuge tubes for the preparation of plasma or may be allowed to clot and 

 the serum centrifuged so as to be absolutely clear. The substratum is the 

 material to be digested. As a rule, the tissue is cleared of connective tissue 

 in so far as possible, is perfused with salt solution and subsequently washed 

 several times with distilled water until it is absolutely free from blood. It is 

 then placed in a suitable container, coagulated by boiling, and repeatedly 

 washed with boiling water until the fluid gives no ninhydrin test. It is then 



