Patent Report

proteins are produced. The HBV antigens expressed by the mammal ian cells can now be prepared on a large-scale and are useful for producing vaccines and diagnostic reagents. A preferred vector, pXR11G, consists of pBR322 deficient in a region toxic to mammal ian cells, plus the SV40 DNA replication origin inserted at the EcoRI site. This is cleaved with e.g. BamHI and incubated with a fragment of the HBV genome. 2-4 HBV fragments can be inserted/plasmid fragment. The product is used to transform mouse LTK-or COS cells in the presence of a large excess of DNA coding for thymidine-kinase. The BamHI fragment of pXRI1G and the BamHI fragment of HBV DNA were incubated in the presence of T4 ligase and the reaction mixture was used to transform E. coil chi 1776 cells. The transformants were selected and plasmids extracted from the isolated clones, one of which contained pSHB3. 058-84

Antigenic herpes simplex polypeptide production by culturing microorganisms containing a vector including a viral DNA fragment

MoL Genetics Eur 101-655, 29 February 1984

Production of polypeptides having an immunoreactive and antigenic determinant of a herpes simplex virus (HSV) glycoprotein comprises cultivating a unicellular organic containing a recombinant vector comprising a DNA sequence coding for the polypeptide (A). (A) Is recovered from the culture. (A) Are useful for producing vaccines against HSV. Various fragments of the Us region of HSV-I DNA were incorporated into pBR322; 1 of the recombinants (pRWF6) contained the entire gD-1 gene. This plasmid was cleaved with SacI and a fragment was ligated with pBR322 to give plasmid pSC30-4. A fragment was isolated from this with PvuIl and SacI, and this was ligated with the SmaI/Sacl fragment from pJS413. The recom- binant plasmids were used to transform Escherichia coli NF 1829 and 1 of the transformants contained plasmid pEH25. This produced a product of MW46 000 containing 23 amino acids of the host protein and 342 amino acids of the gD-I protein. The fusion product was immunoprecipitated. 059-84

Influenza vaccine preparation by extraction of live influenza virus with ether and ethanol mixture

Merck-USA US 4,431,633; 14 February 1984

Production of an influenza vaccine comprises extraction of live influenza virus with a mixture of 98-90% ether and 2-10% ethanol. With the extraction, the virus is detoxified without lowering of immunogenic potency and residual solvent in the vaccine can easily be removed. The extraction is preferably at 5- 25°C. The ether l-ayer is discarded and the inactivated virus is recovered from the aqueous phase. Typically a mixture of 0.5-5 vol of ether and 0.02-0.1 vol of ethanol is used per vol of live aqueous influenza virus. The live virus is concentrated by sedimentation from the egg allantoic fluid in which it is grown and then dialysed against phosphate-buffered saline. The extraction is then performed. 060-84

Cat leukemia associated neoantigen production by culturing infected cells in serum free medium concentration of supernatant and inhibition of protease

Ohio State Univ. US 4,434,157:28 February 1984

Production of cat leukemia associated neoantigen (A), free of virus and cells, comprises first growing cells infected with the virus in a serum-containing growth medium. The cells are transferred to, and grown in, a serum-free medium so that (A) is shed from the cells. The cultured cells are separated from the medium, preferably by centrifugation, and the volume of cell and serum free medium is then reduced to (A)-rich concentrate. The concentrate is then treated to inhibit protease enzyme. Preferably the cat lymphoblastoid cells are cultured and the cell-free (A) containing medium is concentrated by lyophilization, ultra- filtration or especially continuous flow molecular filtration using a filter of exclusion size about MW 10000. The (A) concentrate is useful for making cat leukemia virus vaccine. 061-84

226 Vaccine, Vol. 2, September 1984