How should mice be properly frozen? - briefly
Mice must be anesthetized, transferred into labeled cryovials with suitable cryoprotectant, and moved immediately to a -80 °C freezer or liquid‑nitrogen vapor phase for long‑term storage. Keep the containers sealed, monitor temperature continuously, and prevent any thaw‑refreeze cycles to maintain tissue integrity.
How should mice be properly frozen? - in detail
Freezing laboratory mice demands a controlled, reproducible procedure that preserves tissue integrity and genetic material for future analysis. The process begins with proper animal selection and humane euthanasia, followed by immediate preparation for cryopreservation, precise temperature management, and systematic storage.
First, verify that the subjects are disease‑free and have been acclimated to the facility. Perform euthanasia using an approved method such as CO₂ inhalation or overdose of anesthetic agents, ensuring rapid loss of consciousness to minimize stress‑induced metabolic changes. Immediately after death, remove excess fur and cleanse the carcass with sterile saline to eliminate surface contaminants.
Next, follow a step‑by‑step freezing protocol:
- Pre‑cooling – Place the mouse in a sealed, cryogenic‑grade bag containing a suitable cryoprotectant (e.g., 10 % dimethyl sulfoxide in phosphate‑buffered saline). Submerge the bag in a 4 °C refrigerator for 30 minutes to equilibrate temperature.
- Controlled rate cooling – Transfer the bag to a programmable freezer or a passive cooling container (e.g., isopropanol‑filled Mr. Frosty) set to achieve a cooling rate of approximately –1 °C per minute until the temperature reaches –80 °C.
- Long‑term storage – Move the frozen specimen to a vapor‑phase liquid nitrogen system or an ultra‑low‑temperature freezer maintained at –150 °C or lower. Ensure that the storage unit provides continuous temperature monitoring and alarm systems for deviations.
Maintain records for each specimen, including strain, age, sex, date of euthanasia, cryoprotectant composition, and exact cooling parameters. Periodically retrieve test samples to assess viability of DNA, RNA, and protein extracts; adjust the protocol if degradation is detected.
Adhering to these steps guarantees that frozen mice retain the biological fidelity required for downstream molecular, histological, or genetic investigations.