VIROLOGICAL METHODS 269 



times, without propagation, the virus will disappear. The two methods 

 can be combined by noting the titer of some function upon each serial 

 transfer. If it remains constant or increases, propagation must be 

 taking place. Functional end points vary with the virus under study. 

 In addition to those mentioned under embryonated egg culture, cyto- 

 pathogenic effect may be followed. This will apply wherein the virus 

 can produce inclusion bodies or cellular changes in the host cells. Need- 

 less to say the cells from the control cultures should have their histology 

 scrutinized under the same conditions as the experimental. Every 

 harvest should be checked for bacterial sterility. 



HEMAGGLUTINATION 



A number of viruses will agglutinate red blood cells. Since a given 

 amount of a given concentration of red cells will be agglutinated under 

 standard conditions by a minimal amount of virus, it is possible to titrate 

 this function. Although the hemagglutination test has many modifica- 

 tions, one of the simplest to perform and most likely to give reproducible 

 results in the hands of the inexperienced worker is the pattern test 

 (Salk, 1944). 



Materials. 



1. Serology rack capable of holding 20 Wassermann-size tubes. A piece 

 of white paper may be placed on top of the bottom of the rack, or 

 better still, the metal bottom may be replaced by a transparent 

 material such as a i-^-in. -thick Lucite slab. The latter rack is then 

 set on a white background or on frosted glass with a hght source 

 beneath. 



2. Wassermann tubes that have been carefully cleaned and rinsed free 

 from any cleaning reagent. They should be clear and dry. 



3. Saline solution made from redistilled water and containing 0.85 per 

 cent of sodium chloride may be used as a viral diluent. Under special 

 circumstances a phosphate buffer of 0.1 M and pH 7.0 may be used. 

 This is particularly true if the saline solution is allowed to become 

 very acid while in use. 



4. Red blood cells from adult roosters or 0-type red cells from man 

 serve well, although erythrocytes from some other animals may be 

 used. Where chickens are used, blood may be collected from the wing 

 or jugular veins in citrate solution. After the blood is centrifuged, the 

 plasma and buffy coat are removed and the red cells washed three 

 times. During the last centrifugation at 1,200 rpm for 10 min a cali- 

 brated centrifuge tube may be used in order to make the packed cells 

 into a 3 per cent suspension in physiologic saline. The suspension 



