Topic 8: Tissue Culture Techniques

Tissue culture is a laboratory technique used to grow, maintain, or propagate cells, tissues, or organs in an artificial medium under sterile conditions. It is widely used in research and applications across virology, microbiology, medicine, and agriculture.

Types of Tissue Culture

1. Cell Culture: Growing isolated cells from tissues in vitro.
   • Primary Cell Culture: Cells taken directly from a tissue and grown.
   • Continuous Cell Lines: Immortalized cells capable of indefinite growth (e.g., HeLa, Vero).
2. Organ Culture: Maintaining the architecture of an organ or tissue for research or testing.
3. Explants Culture: Small tissue sections used to initiate cultures.
4. Callus Culture: Used in plant tissue culture to grow undifferentiated plant cells.
5. Protoplast Culture: Growing cells without their cell walls (used in plant research).

General Steps in Tissue Culture

1. Preparation
   • Sterile Environment: Aseptic conditions are critical to prevent contamination.
   • Media Selection: Choose an appropriate nutrient-rich medium like DMEM, RPMI, or MS (Murashige and Skoog) for plants.
2. Tissue Collection
   • Excise tissue from the source organism under sterile conditions.
3. Disinfection
   • Surface sterilize the tissue with solutions like 70% ethanol or sodium hypochlorite.
4. Inoculation
   • Place tissue or cells into culture media in sterile containers.
5. Incubation
   • Maintain cultures under controlled temperature, humidity, and light (if needed).
6. Observation and Maintenance
   • Regularly monitor for growth and contamination.
   • Subculture to fresh media when necessary.

Tools and Equipment

• Laminar Flow Hood: Ensures sterility during procedures.
• Incubator: Maintains optimal temperature and CO₂ levels.
• Autoclave: Sterilizes equipment and media.
• Microscope: Observes cellular growth and morphology.

Applications of Tissue Culture

1. Virology
   • Virus isolation and replication studies.
   • Vaccine development and production.
2. Cancer Research
   • Studying tumor cells and drug efficacy.
3. Biotechnology
   • Genetic engineering and cloning.
4. Pharmacology
   • Drug testing and development.
5. Agriculture
   • Producing disease-free plants and somatic hybrids.