582 IMMUNITY 



stances which when present in the bacterial bodies give rise to 

 agglutination, on the addition of the anti-serum, produce a 

 precipitate when free in a fluid. To test the reaction it is 

 accordingly necessary to have as far as possible the substance of 

 the bacteria in solution, and for this purpose there have been 

 introduced various methods, of which the two following may be 

 given : 



(a) It is well known that in an old bouillon culture the bacteria undergo 

 disintegration and their constituents go into solution. Accordingly, if 

 such a culture which has been kept in the incubator for several weeks be 

 filtered through a porcelain filter, the filtrate will contain the interacting 

 substance or precipitinogen. 



(&) The growth from a recent agar culture is scraped off and suspended 

 in normal salt solution, the mixture is made feebly alkaline with soda 

 solution and boiled for a few minutes. The mixture is then neutralised, 

 when a precipitate forms, and is filtered through filter-paper ; the filtrate 

 contains the precipitinogen. 



The test is carried out by placing in a number of small test- 

 tubes a given amount of the bacterial nitrate along with varying 

 quantities of the homologous anti-serum. (The latter may be 

 obtained in the usual way by the repeated injection of dead 

 cultures or of bacterial nitrate.) As the precipitate forms 

 slowly the tubes should be placed in the incubator for twenty- 

 four hours, *5 per cent, carbolic acid being added to prevent the 

 growth of bacteria. This precipitin reaction has now been 

 observed in a great many bacterial diseases when the patient's 

 serum is added to the corresponding bacterial nitrate, and has 

 even been applied by some observers as a means of diagnosis. 

 It is, however, less delicate and more restricted in its application 

 than the agglutination methods. 



Serum Precipitins. This subject does not strictly belong to bacteri- 

 ology, but the general phenomena are so closely allied to those just 

 described that some reference may be made to it. When the serum of 

 an animal is injected in repeated doses into another animal of different 

 species, after the type of an immunisation, there appears in the serum of 

 the animal treated a substance called precipitin, which causes a cloudi- 

 ness or precipitate when added to the serum (precipitinogen) used. (In 

 the case of rabbits, doses of 3 to 4 c.c. of the serum may be injected intra- 

 peritoneally at intervals of four to five days, a precipitin usually appearing 

 at the end of about three weeks.) The reaction, which is a very delicate 

 one, is conveniently observed by adding a given amount of the anti- 

 serum, say '05 c.c., to varying amounts of the homologous serum '1, '01, 

 etc., c.c., in a series of small test-tubes, the volume being then made up 

 with salt solution to 1 c.c. In this way a definite reaction may be 

 observed with '001 c.c. of the serum or even less. Here again zone 

 phenomena, as in the case of agglutination, are met with. If the anti- 

 serum be heated to a temperature of 75 C. for some time it acquires 



