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Cryopreservation of human granulocytes in liquid nitrogen
Abstract:
Human granulocytes (PMNL) were successfully cryopreserved for up to 14 months. The PMNL (1-2 X 10(7)/ml) were stored in 2-ml ampoules in the gas phase of liquid nitrogen at a temperature between -160 degrees C and -196 degrees C using dimethylsulphoxide (DMSO 10%) as cryoprotectant. Morphology and phagocytic and bactericidal capacity were best preserved by adding fetal calf serum to the freezing mixture, by using an interrupted cooling process, by washing the thawed PMNL in fresh freeze-dried plasma, and centrifuging at 600 g for no more than two minutes. Careful post-thaw handling of the cells was an important factor in preserving function. These preliminary studies indicate that useful numbers of PMNL can be recovered in a functional state after storage for long periods in liquid nitrogen.
Insights
Human granulocytes, also known as polymorphonuclear leukocytes (PMNL), can be successfully cryopreserved for over a year. Optimal methods preserve their morphology and crucial functions for potential therapeutic applications.
Area of Science:
- Immunology
- Cell Biology
- Cryobiology
Background:
- Human granulocytes (polymorphonuclear leukocytes, PMNL) are critical immune cells.
- Effective cryopreservation of PMNL is essential for their long-term storage and potential therapeutic use.
Purpose of the Study:
- To investigate the feasibility of cryopreserving human granulocytes (PMNL).
- To determine optimal conditions for preserving PMNL morphology and function after long-term storage.
Main Methods:
- PMNL were cryopreserved in the gas phase of liquid nitrogen (-160°C to -196°C) using 10% dimethyl sulfoxide (DMSO).
- Cryopreservation protocols included variations in cooling processes, cryoprotectant additives (fetal calf serum), and post-thaw washing procedures (fresh freeze-dried plasma, centrifugation).
Main Results:
- Successful cryopreservation of PMNL for up to 14 months was achieved.
- Preservation of PMNL morphology, phagocytic capacity, and bactericidal capacity was optimized by specific methods.
- Key factors included adding fetal calf serum, using interrupted cooling, washing with plasma, and controlled centrifugation post-thaw.
Conclusions:
- Human granulocytes (PMNL) can be cryopreserved for extended periods (up to 14 months) in liquid nitrogen.
- Careful optimization of cryopreservation and post-thaw handling is crucial for maintaining PMNL viability and function.
- These findings suggest the potential for storing functional PMNL for future clinical applications.