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complement. The anti-complementary fraction was shown to be antigenica 1 ly 
related to _E. col i 1 ipopolysaccharide or endotoxin, and had essentially no 
anti-tumor activity. 
I would like to summarize briefly the experience of Eli Lilly and Company 
with L-asparaginase from _E. col i B. We began producing it experimentally on 
a very small scale in 1968, and by July of that year had a preparation which 
we thought would be suitable for clinical evaluation. Some of the safety 
testing of this material which is pertinent to our discussion is shown in 
Table 4. This lot was produced before we learned how to crystallize the 
enzyme and the specific activity was only 132 I.U./mg. I think the 
important data from this table are that the material was essentially 
non-pyrogenic and had no detectable endotoxin within the limits of the 
assay. This material was not used clinically as we were already starting to 
crystallize the enzyne at that time and we opted to go to the clinic only 
with the purified crystallized enzyme (1) as shown in Figure 3. The process 
we finally developed for clinical trial lots of asparaginase is summarized 
in Table 5. It was a straightforward purification procedure consisting 
chiefly of ammonium sulfate and ethanol fractionations followed by two 
crystallizations from ethanol-water mixtures. No Sephadex or ion exchange 
chromatography was used. 
Polyacryl amide gel electrophoretic patterns of asparaginase are shown in 
Figure 4. Gel number one shows the ethanol precipitated enzyme from step 4 
of the previous figure. Gel 2 is the pattern of the enzyme immediately 
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