for cytopathic effect for the entire 28 day period. Hemagglutination and 
hemadsorption are performed twice during the assay. 
VI.D.5. In Vivo Virus Assay (Protocol 1514.005002 MA, Inc.): 
This test includes the inoculation of adult mice, suckling mice, guinea pigs 
and embryonated hens’ eggs via yolk sack and allantoic routes of inoculation. All 
animals are observed for clinical signs of viral infection. Suckling mice are 
sacrificed on day 14 for the preparation of a tissue homogenate which is inoculated 
into a new group of suckling mice. Specific Pathogen Free (SPF) eggs are examined 
for viability and the blind passage of fluids into new eggs is performed. 
VI.D.6. Karyology Assay (Protocol 1516.018 MA, Inc.): 
This test is designed to confirm the species identity of cultured cells by means 
of isozyme and cytogenetic analysis. 
VI.D.7. Tumorigenicity Assay (Protocol 1514.001 MA, Inc.): 
This study utilizes athymic nude mice which are inoculated subcutaneously 
with cells. Animals are observed for clinical signs, palpated bi-weekly, and 
submitted for histopathology evaluation of multiple tissues. Any animal which 
shows a regressing nodule is sacrificed and submitted for histopathological 
examination. 
VI.D.8. EBV Probe Assay (Protocol 1516.104 MA, Inc.): 
The purpose of this study is to detect EBV DNA that may be present in the 
test article as determined by Southern hybridization using a labeled DNA probe. 
DNA is extracted from the test article cell pellet and blotted onto membranes. 
Blotted membranes are hybridized to a labeled EBV probe and detected by 
autoradiography. DNA from Namalwa cells are included as a positive control. 
VI.D.9. Cytomegalovirus (Protocol 1514.030 M.A., Inc.): 
The test and control articles are directly inoculated onto cell cultures 
(indicator cells) and examined for 42 days for the appearance of cytopathic effect. 
Additionally, cells are fixed and examined using immunofluorescent techniques. 
VI.D.10. HIV Co-cultivation Assay (Protocol 1516.015001 MA, Inc.): 
This procedure is designed to detect small amounts of retrovirus that may be 
present in the test article. To amplify any virus present, selected Peripheral Blood 
Lymphocytes (PBL) cells are infected, mixed with test article, passaged and 
analyzed for HIV by observing for cytopathic effect (CPE) and production of viral 
antigens by antigen capture ELISA. 
VI.D.ll. Bovine Virus Assay (Protocol 1514.032001 MA, Inc.): 
This study is designed to detect bovine viruses such as Bovine Viral Diarrhea 
Virus (BVD), Infectious Bovine Rhinotracheitis Virus (IBR), Parainfluenza 3 (PI 
3), Bovine Adenovirus (BA), or Bovine Parvovirus (BP) using sensitive indicator 
cells and immunofluorescence and cytopathic effect (CPE) as endpoints. 
Recombinant DNA Research, Volume 17 
[501] 
