every step. He said he believed this was good so that the public 
would know there had been careful evaluation at every step. He 
noted, however, the RAC is not an investigative committee and he 
did not believe the purpose of the RAC was to question whether he 
knew how to do an experiment with retroviruses, whether 
Dr. Blaese knew how to grow T cells, or whether Dr. Rosenberg 
knew how to treat cancer, but that its function was to protect 
the public safety. The presentation, he said, was intended to 
demonstrate that there are extensive data showing both the 
feasibility of the protocol as well as its safety. He added that 
further information was available to members of the RAC in the 
IND application being submitted to the FDA. 
Dr. Anderson said data had been presented to demonstrate the 
answers to the first three questions that he identified earlier. 
The fourth question was whether or not the protocol represented a 
significant new threat to the patient. To answer this, he 
proceeded to offer quantitative data to show that the TIL contain 
no infectious viral particles. 
Dr. Anderson said he employed the S+/L- (sarcoma positive/ 
leukemia negative) assay, using a supernatant from 4070A, a wild 
type murine amphotropic virus to detect infectious viral 
particles. This assay can be quantitated to detect the presence 
of a single infectious viral particle in supernatant diluted by 
3x10'* or 3.3x10 s . He noted that a paper describing this 
procedure was being submitted for publication in Virological 
Methods. A 3T3 amplification test is performed in order to 
double-check the procedure. In this assay, various supernatants 
are plated directly onto NIH 3T3 cells, which are very permissive 
for replication of retrovirus, then grown for a week until the 
whole plate is covered, and examined for foci at one and three 
weeks . 
Dr. Anderson said experiments were performed in which the 
supernatant was intentionally contaminated in order to titer 
infectious viral particles. He showed data from three patients 
demonstrating the absence of helper virus in transduced TIL. 
When samples were intentionally contaminated, usually only about 
6 foci could be detected. The highest number of foci that could 
be detected was 40 or a titer of 2-5xlO s causing him to conclude 
that N2-transduced human TIL are generally virus-free. Even when 
intentionally infected, human TIL support murine amphotropic 
virus only at an extremely low titer and some human TIL don't 
support it at all. He noted that the bulk of supporting data for 
this could be found in the IND which was submitted to the FDA. 
Dr. Anderson presented results of polymerase chain reaction (PCR) 
assays looking for DNA from the packaging cell line. Using a 
series of primers, marked and unmarked TIL from 3 patients were 
analyzed. Although PCR is difficult to quantitate, it is 
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