VIROLOGICAL METHODS 255 



and is best determined by comparing the experimental animals with 

 appropriate controls. Quite commonly experimental animals die within 

 24 hr after exhibiting the first neurological signs. 



Intracutaneous inoculations produce a variety of lesions. These should 

 be described accurately as to character and time of appearance. Ery- 

 thema, edema, size, configuration, and differentiation should be deter- 

 mined. Is the lesion macular, papular, vesicular, pustular, or ulcerative? 

 What is the duration of the lesion and each successive change? Does it 

 heal with or without scarring? It is Avell to measure the lesion. Histo- 

 logical study of the involved areas at various time intervals is advisable 

 in some instances. 



A given property of a virus does not necessarily correlate with another 

 property of that virus. To assume that a certain quantitative increase 

 in any one property, for example, hemagglutinin potency, is a direct 

 indication of other functions, such as infectivity titer, is not only unwar- 

 ranted but may lead to considerable error. Under certain standardized 

 conditions direct relationships may hold, but too much faith should not 

 be extended in this direction. The time the virus is harvested may 

 influence the various propensities of the virus, for it is known that some 

 functions may increase before others. 



It is not particularly difficult to follow the increase of a well-adapted 

 virus in animals. Following the progress of a freshly isolated strain or 

 determining if a virus has been isolated may be very difficult. An 

 appropriate number of days ma}^ elapse after the inoculation of a labora- 

 tory animal with material suspected of containing a virus without any 

 evidence of infection. The experienced investigator does not accept this 

 as a failure but harvests the tissue that should be involved and prepares 

 it for passage to new animals. This procedure has been referred to 

 variously as "passage by faith" and "blind passage." The object is to 

 cause an inapparent infection to become an apparent one. After several 

 passages one concludes that he has failed or is rewarded by noting some 

 evidence of infection. Serial passage of some tissues, for example lungs, 

 at too short an interval encourages bacterial pneumonia. If a 4-day 

 interval is used, this complication usuall}^ does not occur. Too long an 

 interval between passages encourages antibody production and results in 

 defeat for this reason. 



Inapparent infections may be made obvious sometimes by other 

 methods. Although an infection may not have progressed to the state 

 of recognition, it may have induced antibody formation. The tissue or 

 organ used for serial passage may also be examined for the presence of 

 specific antigen. The appearance of this antigen after several passages 

 should encourage continued passage. 



The repeated passage of viruses or rickettsiae through experimental 



