PRESERVATION OF MICROORGANISMS

LABORATORY PROCEDURES FOR MICROORGANISMS

REFERENCE NO.: M/1998/3.00

TITLE: PRESERVATION OF MICROORGANISMS

PRESERVATION

Microorganisms require special preservation methods in order to ensure optimal long-term viability and genetic stability.

In general each preservation method can be assigned to one of the following groups:

1. Metabolically inactive preservation techniques (CABRI accepted methods)

1.1. Cryopreservation

1.1.1. Freezing and low temperature storage in or above liquid nitrogen

1.1.2. Freezing and low temperature storage below -70°C

1.2. Drying

1.2.1. Preservation by shelf freeze-drying

1.2.2. Preservation by spin freeze-drying

1.2.3. Preservation by liquid drying (L-drying)

1.2.4. Preservation by vacuum drying

2. Metabolically active methods

2.1. Periodic transfer on agar or in liquid medium

2.2. Keeping agar cultures under mineral oil

Maintenance of strains by metabolically active methods should be used only in case a strain cannot be preserved by one of the metabolically inactive methods, or in addition to one of these methods.

Flow charts for preservation methods are given in M/1998/3.00 Appendix 1 and M/1998/3.00 Appendix 2. Examples of preservation methods used in CABRI collections are summarised in M/1998/3.00 Appendix 5.

For the protection of cells against damage during freezing and (freeze)-drying as well as during storage microorganisms are normally suspended in a protective suspension medium. Some examples are described in M/1998/3.00 Appendix 3. M/1998/3.00 Appendix 4 gives an example of a protocol form for preparing protective media.

The preservation methods used in the collections may differ. Each collection must maintain detailed protocols of the applied preservation methods and their application to specific groups of microorganisms. Examples of them are given in M/1998/3.00 Appendix 5.01 to 5.13. If available, registration form for freeze-drying process per batch (e.g. vacuum, product temperature, shelf temperature, condenser temperature, time) should be filed.

QUALITY CONTROL AFTER PRESERVATION

After every preservation of a microbial strain controls are necessary. At least viability and purity, and where appropriate, the identity of the preserved culture have to be checked immediately after preservation. Details of these controls are inserted in the protocols of the preservation. If available a registration form for the freeze-drying process per batch (e.g. vacuum, product temperature, shelf temperature, condenser temperature, time) is filed.

Any remark on the viability or properties of a batch has to be archived and remain available to compare with future controls.

Methods for the safe opening of glass capillaries are given in M/1998/3.00 Appendix 5.01. Methods for the safe opening of ampules of freeze-dried cultures are given in M/1998/3.00 Appendix 5.14.

Guidelines prepared for CABRI by DSMZ, CBS and BCCM, 17 May 1998
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