22 MASS. EXPERIMENT STATION BULLETIN 323 



Discussion 



From results obtained in this investigation, the whole blood test, as a diagnostic 

 means of detecting infected pullorum-disease carriers, does not appear to be so 

 efficient as the standard tube method. Antigen and whole blood measurements 

 demonstrated that the quantities of antigen and of whole blood might vary con- 

 siderably, and that the whole blood-antigen dilution was by no means constant. 

 This variation in the whole blood-antigen dilution had a perceptible influence 

 on the degree of agglutination and was especially significant in birds that pos- 

 sessed a low agglutination titre. Failure of such birds to react might be due to 

 the whole blood-antigen dilution being beyond the agglutination titre. While a 

 definite amount of blood appeared essential to obtain a reaction, yet a greater 

 amount of blood did not seem to increase the intensity and rapidity of the agglu- 

 tination. Equal amounts of blood and antigen appeared to give the best results, 

 while amounts of blood greater than that of antigen seemed to form a colored 

 mixture in which the clumping was observed with difficulty. 



The whole blood and antigen dilution does not appear to be the only factor 

 which influences the accuracy of the whole blood method. While the exact in- 

 fluence which uniform and constant incubation temperature exerted on the 

 agglutination reaction was not definitely determined, it was observed that a 

 controlled heating device was necessary to avoid extremes in temperature which 

 would lead to an unsatisfactory test. The temperature range in which agglutina- 

 tion is not hindered can only be determined when other steps in the test are kept 

 uniform and constant. 



An adequate source of light is essential for accurate interpretation of the 

 agglutination reactions. With a suitable light the clumping will appear distinctly 

 and plainly visible, even when the clumps are few in number and small in size. 

 With insufficient light the weak reactions may be completely overlooked. 



Interference with the tests due to foreign material, such as dust particles and 

 feathers, can be largely overcome by using a covered test plate. The glass test plate 

 marked off in one-inch squares with a diamond pencil prevents the whole blood - 

 antigen tests from spreading and running together. Rotating the test plate 

 appears to hasten the clumping of the cells as well as to bring about a grouping 

 of the clumps. The latter is especially true in strong reactions. 



To what extent humidity, air currents, and other environmental factors in- 

 fluence the agglutination reaction remains a question. The fact that there has 

 been no satisfactory investigation of the influence which any of the apparently 

 important factors exert upon the agglutination reaction, makes it impossible to 

 ascribe to the different factors their full significance in the whole blood agglutina- 

 tion phenomena. 



Since it is impossible to produce on the poultry farm conditions which simulate 

 those in the laboratory, it appears reasonable that the quality of the test con- 

 ducted in the field would not be equal to that conducted in the laboratory. The 

 rate of speed at which the testing is conducted appears to have a marked in- 

 fluence. Some persons are prone to sacrifice accuracy for speed. Such a service 

 may be satisfactory to certain flock owners, but in an eradication program success 

 is based upon accurate and reliable testing results. Sufficient time should be taken 

 to measure the blood and antigen as accurately as possible, to mix them thoroughly 

 and to incubate them for a satisfactory period. The length of the incubation period 

 considered necessary for agglutination to occur varies with different investigators. 



