USSR’s Biological Warfare: Tissue Culture Technology

Possible Deadly Human Virus Research?

Soviets have been considerably interested in the cultivation and use of human diploid cells. Dr. R. I. Rapoport, MNllVP, isolated 13 strains of diploid cells from human embryo lung tissue and determined the stability of the diploid cells in various media. A long-term study of one line (L-10) showed that the best results were obtained with Eagle's mediwn No. IX, which contained 10% bovine serum, the diploid complement of chromosomes was preserved, and the line was devoid of oncogenic (tumor-producing) effects. The L-10 cell line was also- sensitive to measles, tick-borne encephalitis (TBE), and adenoviruses.

A semiautomatic system for propagation of tissue cells was described by L. N. Mishin, Institute of Virology imeni Ivanovskiy (see figure 2). nie concentration of air or co2 in a gas mixture-;as automatically adjusted to maintain a preset level of CO:z in a sodium bicarbonate-buffered medium. Mixing was provided by a float-type blade mounted inside the culture vessel and driven by a magnetic stirrer. nie use of a magnetically driven blade eliminated the need for a shaft seal and reduced contamination problems. The apparatus, operated semi-continuously for several months, maintained good culture conditions. Infectivity studies using VEE virus and chick embryo fibroblast cells showed that the device could be used to propagate viruses. A titer of 2.1 x 109 virus particles per milliliter was obtained at a cell concentration of 2 x 106 cells per milliliter.

Work on automated tissue cell culture systems indicates that the Soviets have successfully copied techniques previously reported and have incorporated modifications which contribute to the simplicity and dependability of operation. 11te method of continuous cell cultivation is well suited to the industrial production of cell cultures, antigens. viruses, and nucleic acids in mass quantities. The method is more advantageous than ordinary systems for producing viruses and may make it possible to obtain large concentrations of viruses for the solution of many theoretical and practical problems. In addition, the cultivation of TBE, VEE, and adenovi'ruses shows much promise.

TISSUE CULTURE IN VIROLOGY

Soviet scientists have employed many of the established cell lines and have initiated many other cell cultures for virus investigations. The intensive interest shown by Soviet researchers and the wide distribution of work with human diploid cells indicate a concentrated effort to thoroughly explore the use of diploid cells in virology.

Soviet researchers have employed monolayer tissue cell cultures to produce vaccines, and the method could be used to grow cells for the production of viral agents. The system is impractical, however, because the contents of large numbers of flasks must be pooled, and the procedure is cumbersome, time consuming, and laden with contamination problems.

Institute of Virology imeni Ivanovskiy, used a similar system to produce rabies virus in Syrian hamster kidney fibroblasts (BHK-21 cell line). Bottles with capacities ranging from 1 to 20 liters were used, with the volume of medium being about one-twentieth that of the bottle capacity. Inocula were added to give an initial concentration of 4 to 5 X la5 cells per milliliter. A special rack with different-sized rollers was designed to rotate bottles, regardless of size, at 0-. 7 to 0, 8 rpm. Cells began to attach to the glass surface 2 hours after inoculation. Most.of the cells were attached in 12 to 18 hours, and a complete monolayer was formed after 24 to 48 hours.

Primary infection of the cells was accomplished after 24 to 48 hours of cultivation by introducing a 100/o suspension of Brain cells infected with rabies virus at one-fifth the volume of the medium. Subsequent infections were made by subculturing methods or by mixing infected cells with uninfected cells in fresh medium

Sources:

The Black Vault

Defense Intelligence Agency