The “Stained Slide” Microscopic Agglutination Test

Abstract

The importance of obtaining an early diagnosis of the type of infecting pneumococcus, especially in cases of lobar pneumonia to be treated with serum, has resulted in the development of many methods. The microscopic agglutination test just described, since it required very few organisms, was expected to reduce the time for a determination of type. Attempts to type the organisms directly from the sputum were unsuccessful. The fact that in the microscopic test the sputum was exposed to the immune serum for a very short time only, was at first thought to be responsible for the negative results. Small amounts of sputum were accordingly mixed with varying amounts of serum, and incubated in sealed capillary tubes at 37° C. for different periods of time; these mixtures were subsequently smeared and stained. The unsuccessful results are due possibly to the presence of some interfering factor or unfavorable physical condition in the substance in which the pneumococci are coughed up. Most of the sputa injected into the mouse's peritoneum are digested within 2 hours, and in 3-4 hours, sufficient organisms may be found to carry out a microscopic typing test. Broth may be used as a culture medium in the absence of mice, but the mouse is preferred because its peritoneum is somewhat selective for pneumococci, and because results can be obtained in a shorter time.

The procedure for microscopic typing is as follows: One cc. of a fresh sample of sputum is injected intraperitoneally into a mouse. 3-4 hours after injection some of the peritoneal fluid is obtained by capillary puncture. A glass slide is marked off into 4 parts, and a minute drop of the peritoneal fluid is expelled onto each one of the 4 partitions. The first is smeared with saline for control, and the others with a loopful of a 1-10 dilution of type I, of type II, and of type III, diagnostic serums respectively. This dilution of serum is chosen to largely eliminate group agglutinins.