1.Laboratory Design

Perhaps one of the most under-rated aspects of tissue culture is the need to design the facility to ensure that good quality material is produced in a safe and efficient manner. Most tissue culture is undertaken in laboratories that have been adapted for the purpose and in conditions that are not ideal. However, as long as a few basic guidelines are adopted this should not compromise the work.

There are several aspects to the design of good tissue culture facilities. Ideally work should be conducted in a single use facility which, if at all possible, should be separated into an area reserved for handling newly received material (quarantine area) and an area for material which is known to be free of contaminants (main tissue culture facility). If this is not possible work should be separated by time with all manipulations on clean material being completed prior to manipulations involving the ‘quarantine material’. Different incubators should also be designated. In addition, the work surfaces should be thoroughly cleaned between activities. All new material should be handled as ‘quarantine material’ until it has been shown to be free of contaminants such as bacteria, fungi and particularly mycoplasma. Conducting tissue culture in a shared facility requires considerable planning and it is essential that a good technique is used throughout to minimize the risk of contamination occurring.

For most cell lines the laboratory should be designated to at least Category 2 based on the Advisory Committee on Dangerous Pathogens (ACDP) guidelines (ACDP, 1985) *. However, the precise category required is dependent upon the cell line and the nature of the work proposed. The guidelines make recommendations regarding the laboratory environment including lighting, heating, the type of work surfaces and flooring and provision of hand washing facilities. In addition it is recommended that laboratories should be run at air pressures that are negative to corridors to contain any risks within the laboratory.

*Advisory Committee on Dangerous Pathogens (1985) Categorization of Biological Agents According to Hazard and Categories of Containment, 4th edition, HSE books, Sudbury, UK.

2.Microbiological Safety Cabinets

A microbiological safety cabinet is probably the most important piece of equipment since, when operated correctly, it will provide a clean working environment for the product, whilst protecting the operator from aerosols. In these cabinets operator and/or product protection is provided through the use of HEPA (high efficiency particulate air) filters. The level of containment provided varies according to the class of cabinet used. Cabinets may be ducted to atmosphere or re-circulated through a second HEPA filter before passing to atmosphere.

Environmental monitoring with Tryptose Soya Broth agar settle plates inside the cabinet for a minimum of four hours should be a good indicator of how clean a cabinet is. There should be no growth of bacteria or fungi on such plates.

In most cases a class II cabinet is adequate for animal cell culture. However each study must be assessed for its hazard risk and it is possible that additional factors, such as a known virus infection or an uncertain provenance, may require a higher level of containment.