Preperation of Cell Line_animal cell culture.pptx

Preperation of Cell Line_animal cell culture

1. Preparation of Cell Line

2. Cell Line Cell Line is a propagated culture after the first subculture Derived from primary culture Might contain several cell lineages of either similar or distant phenotypes There are around 4000 different mammalian cell lines available- both from human and non-human origin HeLa : adult human cervical epithelial cell WI38: human embryonic lung fibroblast Caco2: adult human colon epithelial MCF7: Adult human breast tumor epithelial Vero: monkey kidney fibroblast

3. Vero MCF7

4. Common Cell Types Based on Morphology Fibroblast - bipolar or multipolar, have elongated shapes, and grow attached to a substrate Epithelial - polygonal in shape with more regular dimensions, and grow attached to a substrate in discrete patches Endothelial - Endothelial cells of all vascular beds form a single cell layer  in vivo  which is exposed to blood on the lumenal side and to interstitial fluid and extracellular matrix on the ablumenal side Neuronal - exist in different shapes and sizes, but they can roughly be divided into two basic morphological categories,  type I  with long axons used to move signals over long distances and  type II  without axons. A typical neuron projects cellular extensions with many branches from the cell body, which is referred to as a dendritic tree Lymphocyte - spherical in shape and usually grown in suspension without attaching to a surface

5. Sources of Cell There are two methods for obtaining cells: from a cell bank, or by isolating cells from donor tissue

6. Animal Cell Bank/Collection DSMZ- German Collection of Microorganisms and Cell Cultures European Collection of Animal Cell Cultures American Type Culture Collection (ATCC) Japanese Cancer Resources Bank

7. Steps to Obtain a Cell Line from Donor Tissue Sample Tissue Dissection and/or disaggregation Primary culture Subculture Cell Line Cell strain

8. Primary Culture A primary culture is that stage of culture after isolation of cells but before the first subculture Features of primary culture Initially heterogeneous Resembles tissue characteristics and thus closer to an in vivo condition compared to cell line Limited life span Variability exists between donors and preparations Relatively slow growth in culture Needs optimized culture conditions and careful handling Not altered (except dissociation from tissue)

9. Types of Primary Culture Adherent -Require attachment for growth, i.e. anchorage-dependent - Derived from tissue or organ Suspension Do not require attachment for growth, i.e. anchorage-independent Isolated from blood system Might be as well as stem cells or transformed cells

10. Initiation of a Primary Culture Acquisition of the sample Isolation of the tissue Dissection and/or disaggregation Culture after seeding

11. Primary Culture

12. Factors in sample selection and tissue isolation Most common tissue explants: Mouse embryo Chick embryo Human biopsy material Medical ethical rules Legislation on experimentation with animals Safety Contamination Transport and storage