ABSTRACT
After it has been proven that the glycosylated and nonglycosylated IFN
γ
have the same biological activity (see Chapter 2, Section 2.3.1), IFN
γ
is now produced mainly by the methods of recombinant DNA technology (genetic engineering). Eukaryotic (Devos et al. 1982) as well as prokaryotic (Gray et al. 1982; Perez et al. 1990) cells may be employed as producers. The use of eukaryotic cells runs the risk of contamination with potentially dangerous viral and oncogenic DNA. That is why prokaryotic producers are preferred. The most widely used microorganism is
Escherichia coli
transformed with a plasmid containing the gene of IFN
γ
. The gene itself can be obtained by either reverse transcription from IFN
γ
mRNA or by a total chemical synthesis. Usually the production cycle of human recombinant IFN
γ
includes the following steps:
1. Transformation of an appropriate strain with an expression plasmid containing the human IFN
γ
gene 2. Fermentation 3. Collection of the bacterial biomass 4. Disintegration of the transformed cells where rIFN
γ
is usually accumulated as inclusion bodies (Figure 5.1)
5. Disolving the inclusion bodies in a denaturing solution 6. Purification of IFN
γ
by chromatography (Figure 5.2) 7. Control of purity, activity, and identity
Figure 5.1
Electron micrographs of
E. coli
sections containing IFN
γ
stored as inclusion bodies (black spherical formations).
