Maintaining cells, particularly in a laboratory setting (cell culture), involves carefully controlling their environment to mimic their natural conditions, thereby promoting their survival and growth. This primarily includes regulating temperature, gaseous environment, and providing necessary nutrients.
Here's a breakdown of the key aspects of cell maintenance:
1. Controlled Environment:
- Temperature: Cells are typically maintained at a specific temperature relevant to their origin. For mammalian cells, this is usually around 37°C, mimicking the body temperature of mammals. Incubators provide precise temperature control.
- Gaseous Environment: The proper balance of gases, especially carbon dioxide (CO2) and oxygen (O2), is crucial. CO2 helps maintain the correct pH of the cell culture medium. The appropriate O2 level depends on the specific cell type. Specialized incubators control these gas mixtures.
- Humidity: High humidity within the incubator prevents the culture medium from evaporating, ensuring consistent osmolarity and nutrient concentrations.
2. Culture Medium:
- Nutrient Supply: Cell culture medium provides the necessary nutrients (amino acids, vitamins, glucose, salts, etc.) for cell survival and proliferation.
- pH Control: The medium contains buffering systems (e.g., bicarbonate) to maintain the physiological pH (typically around 7.2-7.4).
- Sterility: Strict sterile techniques are essential to prevent contamination by bacteria, fungi, or other microorganisms, which can outcompete or kill the cells. This includes using sterile media, cultureware, and working in a laminar flow hood.
3. Culture Vessels:
- Cells are grown in sterile culture vessels, such as flasks, dishes, or multi-well plates, designed for cell culture. The choice of vessel depends on the scale of the experiment and the cell type (e.g., adherent vs. suspension cells).
- Passaging/Subculturing: As cells grow and proliferate, they need to be periodically transferred to new culture vessels with fresh medium to prevent overcrowding and nutrient depletion. This process is called passaging or subculturing.
4. Monitoring and Observation:
- Regular observation of the cells under a microscope is crucial to assess their health, morphology, and confluency (density).
- Cell viability assays may be performed to quantify the number of live cells in the culture.
5. Preventing Contamination:
- Sterile Technique: Performing all cell culture procedures under sterile conditions, typically within a laminar flow hood.
- Antibiotics: While controversial for long-term culture, antibiotics (e.g., penicillin/streptomycin) are often added to the culture medium to prevent bacterial contamination. However, relying on antibiotics can mask poor sterile technique.
- Regular Testing: Periodically testing the cells for contamination (e.g., mycoplasma) is important to ensure the integrity of the culture.
Example:
Imagine you're maintaining human lung epithelial cells (e.g., A549 cell line). You would:
- Grow them in a cell culture flask in a 37°C incubator with 5% CO2.
- Use a specific cell culture medium formulated for lung epithelial cells, supplemented with fetal bovine serum (FBS) as a growth factor.
- Change the medium every 2-3 days.
- Passage the cells when they reach about 80% confluency.
- Regularly check them under a microscope for signs of contamination or abnormal morphology.
Summary:
Maintaining cells in culture requires meticulous control of their environment, provision of appropriate nutrients, and strict adherence to sterile techniques. Careful monitoring and timely interventions (e.g., passaging, medium changes) are essential for ensuring cell health and viability.
