types of animal cell culture

what is cell culture

the removal of cells from an animal or plant and their subsequent growth in a favourable artificial environment

obtained by

  • removal from the tissue directly and disaggregated by enzymatic or mechanical means bfr cultivation

  • derived from cell line/cell strain (alrdy established)

in vivo - inside a living organism

in vitro - outside of a living organism (in a test tube)

reasons for growing animal cells in labs

  • no animal testing

  • research and study

  • cell-based manufacturing (like vaccines and replacement tissues and organs)

  • medical use (cell therapy, in vitro fertilisation, genetic counselling, drug screening & development)

starting animal cell culture

  1. thaw culture medium vessel by gentle agitation in water bath at 37ÂșC/normal growth temp for that cell line (2 mins/until melted)

  2. decontaminate by spraying/dipping 70% ethanol

  3. transfer contents in vial to centrifuge containing 9mL of recommended medium

    - remove cryoprotectant agent by gentle centrifugation (10min at 125xg)

  4. discard supernatant and resuspend in 1 or 2mL of complete growth medium

  5. transfer cell suspension into culture vessel with complete growth medium

  6. mix thoroughly by gentle rocking

  7. examine after 24h

if cells are attached grow in monolayer

  1. aseptically remove all but 5mL to 10mL of the shipping medium (store remaining shipping medium at 4ÂșC

  2. incubate flask at 37ÂșC with 5% CO2 until cells are subcultured

if cells are not attached/growing in suspension

  1. transfer entire contents of flask to centrifuge tube

  2. centrifuge at 125 x g for 5 to 10 min

  3. remove all but 10mL of shipping medium supernatant and resuspend cells. (store leftover at 4ÂșC)

  4. transfer resuspended cells to 25cm2 flask or 75cm2

  5. incubate cells at 37ÂșC with 5% CO2 until cells are subcultured

Primary cell culture

  • refers to the stage of the culture after the cells are isolated from the tissue and proliferated under the appropriate conditions until they occupy all the available substrate (reaching confluence)

  • cells have to be subcultured by transferring them to new vessel with fresh growth medium to provide more room for continued growth

  • originates from animal or human

  • separates into indiv cells by

    • enzymes (trypsin) that digests away sticky proteins that hold the cells together - incubate at 37ÂșC for optimum enzyme activity with shaking

    • mechanical dispersion - forcing tissue fragments through a syringe collecting the cells when a fragment of tissue is dissected and/or pressing fragments of tissue through a series of cell sieves

  • grown from fragments of tissues (dissected animal tissue is chopped into small pieces

cell line

culture is considered a primary culture up to its first subculture; after that its named a cell line

subculturing of adherent cells

  • reduce the cell density so that they have sufficient space to grow

  • maintain cells at log phase

adherent cells

(type of cell line)

cells that need to be attached to a surface in order to grow are adherent cells

  • cells grow as a monolayer, attached to each other

  • each cell looks flattened and spread out

  • nucleus and cytoplasm can be seen

  • usually come from solid tissue (lungs, kidney)

suspension cells

(type of cell line)

cells are suspended (float) in the culture medium. Suspension cells do not need to be attached to a surface in order to grow

  • each cell looks round

  • nucleus and cytoplasm are not easily differentiated

come from blood forming tissues or from adherent cells adapted to suspension culture