Implementing Best Practices and Validation of Cryopreservation Techniques for Microorganisms

Table 3

Validation of fungal cryopreservation protocols.

Criterion	Requirement	Reference
Select optimal growth conditions	To produce healthy material spores, mycelium	[20]
Measure and record baseline data for stability checks	Apply a unique identifier/strain number; select criteria to be measured: Morphological characteristics Sequence ITS region of the genome; Growth rates Photomicrographs; Metabolic data; Genome fingerprinting techniques	[8]
Select the most appropriate preservation protocol	Optimised for organism type	[20]
Select cryoprotectant	Appropriate for the cell type	[8]
Select most appropriate cooling rate	Thermometer calibrated to a standard	[8]
Select most appropriate storage temperature	Temperature below–140°C, monitored and recorded	[21, 22]
Select most appropriate thawing protocol	A rate appropriate to cell type in calibrated and controlled equipment	[8]
Prepare master and distribution stocks	High recovery; No contamination; Authentic: morphology; phenotypic and molecular integrity	[23]
Method validation
Performing blind tests	Central laboratory sends unknown organism to collections with limited data and results after above the process compared	Example of such a system is in the public health laboratories for diagnostics
Reproducibility check	Comparing results of the same method at different times; Comparison of results obtained with different methods; Comparison of results obtained with different operators	[23]
Equipment calibration	All equipment must be regularly serviced and gauges and meters calibrated to recognised standards	[23]
Recording parameters	Daily records of temperature readings of incubators and cryostorage units to ensure that they remain within set parameters	ISO standards