Defensive Ferments 141 



tissue was in the body, it became a test for the determination of 

 the existence of pregnancy. The method required but little in the 

 way of special apparatus or reagents. The chief requirements 

 being small "dialyzing shells" or thimbles, which are made by 

 Schleichter and Schull, and are commercially known as No. 5790. 

 They are procurable through importing agents dealing in laboratory 

 apparatus. These shells must be tested before using, and it is best 

 to test a large number at the same time. Each must be impervious 

 to albumen, but readily permeable to peptones, aminoacids and 

 other cleavage products of protein digestion. 



The shells or " thimbles" are tested thus by Kolmer:* 



They are first soaked in sterile distilled water for half an hour or more, until 

 they are softened. Each then receives about 2.5 cc. of a 5 per cent, solution of 

 egg-albumen in distilled water, thoroughly mixed and free from flakes or shreds. 

 In filling the shell, care should be exercised that none of the albumen solution by 

 any chance falls upon the outside. The shell is then picked up with forceps and 

 transferred to a short tube containing about 20 cc. of sterile distilled water. 

 This tube should be so wide that the column of water is not so deep as the shell is 

 high, and not so broad that the shell is in danger of oversetting. As bacteria 

 may not have been successfully excluded and by multiplying may cause proteo- 

 lytic cleavage of the albumen, it is well to cover the fluid in the thimble and that 

 in the tube outside of it, with a thin layer of toluol. The outer tube is plugged or 

 corked, and the whole is stood in the incubating oven where it is kept at 37C. 

 for sixteen to eighteen hours. At the end of this time, 10 cc. of the water in the 

 outer tube is removed by a pipette, and tested by the biuret reaction to determine 

 whether any albumen has penetrated the thimble. For this purpose the fluid, 

 in a test-tube, receives 2.5 cc. of a 33 per cent, solution of sodium hydroxid and is 

 shaken gently. One cubic centimeter of a 0.2 per cent, cupric sulphate solution 

 is permitted to trickle down the side of the tube and overlie the contents. If a 

 delicate violet is produced at the line of junction of the two liquids, albumen has 

 escaped from the thimble into the water outside. Under such circumstances the 

 thimble is, of course, useless and should be thrown away. ^ If there is any uncer- 

 tainty about the reaction, the tube can be stood away for eight hours or so longer 

 (twenty-four hours in all) and the remaining water subjected to the ninhydrin 

 test (see below). 



The good shells or thimbles are next to be tested for permeability to peptones. 

 Before this they should be carefully washed in running water and boiled for 

 thirty seconds. 



A i per cent, solution of Hochst "silk peptone" is made in distilled water, and 

 of it 2.5 cc. is pipetted into each thimble to be tested, taking care, as before, that 

 none of the solution by accident drops on the outside of the shell. The shell is 

 now placed in the 20 cc. of sterile distilled water in the wide tube such as was 

 used before, covered with toluol and stood in the incubator at 37C. After 

 twenty-four hours, a pipette is thrust through the toluol and 10 cc. of the water 

 taken up. The finger being held over the top of the pipette, the tube is wiped 

 outside with care, so as to get off any toluol, and the fluid then delivered into a 

 test-tube. Here it receives 0.2 cc. of a i per cent, solution of ninhydrin, and is 

 boiled for exactly one minute. If the peptone has dialyzed, a deep blue color 

 develops after standing for a short time. The thimble that permits no transfu- 

 sion of peptone is worthless and should be thrown away. 



The good thimbles are now again thoroughly washed in running water for a 

 minute, or so, and are then transferred to a vessel of sterile distilled water con- 

 taining chloroform to saturation and covered with toluol. 



In making the Abderhalden test it is imperative that the glass- 

 ware used should be chemically clean, that the reagents be pure, 

 that the preparations be kept sterile and that the thimbles and sub- 

 strata should be handled with forceps, not with the fingers. 



* "Infection, Immunity and Specific Therapy." Phila., 1915; p. 253. 



