Summary
Highlights
Cell culture should ideally be performed in a dedicated facility due to potential health threats from cell lines. It's crucial to consult local safety and environmental regulations and conduct risk assessments. The European Collection of Cell Cultures operates laboratories at negative pressure to contain aerosols. Laboratory surfaces, including benchtops, walls, and flooring, should be smooth, easy to clean, waterproof, and chemical-resistant. Coved skirting is recommended for easier cleaning, and windows should be sealed. Storage areas for liquid nitrogen should have crack-resistant floors.
Microbiological safety cabinets (MSCs), also known as MSC Type 2 cabinets or hoods, are vital for cell culture. They provide a clean working environment and protect the operator through airflow, negative pressure, and HEPA filters. Cabinet exhaust can be ducted or recirculated through a second HEPA filter back into the laboratory.
Regular monitoring of cell cultures is essential for tracking growth and health. An inverted phase microscope is required for this, while a standard light microscope is useful for cell counting. Microscopes are precision instruments that require careful handling, should be turned off, and covered when not in use.
Centrifuges are used to rapidly sediment cells into a pellet for transferring them into different media. A small benchtop centrifuge with controlled braking, swing-out buckets, and sealed caps is typically sufficient. Centrifuges should be placed where they are easily accessible for cleaning and maintenance, away from microscopes. It's important to balance tubes carefully, avoid overfilling, and regularly check for corrosion. Cells usually sediment satisfactorily between 80g and 150g, as higher forces can cause damage or agglutination.
Incubators aim to mimic the in vivo cellular environment. Most mammalian cells require 37°C, 95% humidity, and 5% CO2, but other cell types may need different conditions. Regular checks and replenishment of water in the reservoir, independent temperature verification, and CO2 level monitoring are crucial. CO2 cylinder levels should be checked, especially before weekends and holidays. Incubators are prone to bacterial and mold contamination, so regular monitoring and cleaning are necessary.
Water baths are used for thawing frozen cells, heat inactivation of serum, and warming media and reagents. As potential sources of contamination, water baths must be regularly cleaned and replenished with fresh water treated with an appropriate antimicrobial agent.
Most cell culture reagents require refrigeration or freezing. Always check supplier labels and literature for storage instructions. To prevent repeated heating and cooling, aliquot reagents into suitable volumes and concentrations and label them appropriately. Pouring from container to container is generally not good practice, but it is acceptable when aliquoting entire bottles of media or reagents.
A well-equipped laboratory will have a wide range of consumables. Most cell culture flasks, centrifuge tubes, and pipettes are available in single-use sterile packs, ensuring high-quality assurance and eliminating the need for validating cleaning and sterilization procedures.