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    Biological Techniques and Procedures: Cell Culture

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    caitlynangelie
    Cell culture involves growing cells outside the body in carefully controlled conditions. There are three main types of cell cultures: primary cell cultures grown directly from tissue, secondary cell cultures which are subcultures of primary cells, and continuous cell lines that can be cultured indefinitely. Basic cell culture techniques require sterilization and aseptic techniques to maintain pure cultures. Key factors that affect cell growth include nutrients, temperature, pH, oxygen, water, and light.

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    Biological Techniques and Procedures: Cell Culture

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    caitlynangelie
    88 views22 pages
    Cell culture involves growing cells outside the body in carefully controlled conditions. There are three main types of cell cultures: primary cell cultures grown directly from tissue, secondary cell cultures which are subcultures of primary cells, and continuous cell lines that can be cultured indefinitely. Basic cell culture techniques require sterilization and aseptic techniques to maintain pure cultures. Key factors that affect cell growth include nutrients, temperature, pH, oxygen, water, and light.

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    Cell culture involves growing cells outside the body in carefully controlled conditions. There are three main types of cell cultures: primary cell cultures grown directly from tissue, secondary cell cultures which are subcultures of primary cells, and continuous cell lines that can be cultured indefinitely. Basic cell culture techniques require sterilization and aseptic techniques to maintain pure cultures. Key factors that affect cell growth include nutrients, temperature, pH, oxygen, water, and light.

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    Download as pptx, pdf, or txt
    88 views22 pages

    Biological Techniques and Procedures: Cell Culture

    Uploaded by

    caitlynangelie
    Cell culture involves growing cells outside the body in carefully controlled conditions. There are three main types of cell cultures: primary cell cultures grown directly from tissue, secondary cell cultures which are subcultures of primary cells, and continuous cell lines that can be cultured indefinitely. Basic cell culture techniques require sterilization and aseptic techniques to maintain pure cultures. Key factors that affect cell growth include nutrients, temperature, pH, oxygen, water, and light.

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    BIOLOGICAL TECHNIQUES AND

    PROCEDURES

    CELL CULTURE
    What is a Cell Culture?
     the maintenance and
    growth of the cells of
    multicellular organisms
    outside the body in specially
    designed containers and
    under precise conditions of 
    temperature, humidity,
    nutrition, and freedom from
    Three types of cell cultures?
    Primary cell culture :
    This is the maintenance of
    growth of cells
    dissociated from the parental
    tissue (such as
    kidney or liver) using
    mechanical or
    enzymatic methods, in culture
    medium using
    SECONDARY CELL CULTURES
    When a primary culture is sub-cultured, it
    becomes known as secondary culture or
    cell line.
    Subculture (or passage) refers to the
    transfer of cells from one culture vessel to
    another culture vessel.
    This is periodically required to provide
    fresh nutrients and growing space for
    continuously growing cell lines. The
    process involves removing the growth
    media and disassociating the adhered
    cells (usually enzymatically).
    Cell Line
     A cell line or cell strain may be
    finite or continuous depending
    upon whether it has limited culture
    life span or it is immortal in culture.
    On the basis of the life span of
    culture, the cell lines are
    categorized into two types:
    Finite cell lines :
     The cell lines which have a limited life
    span and go through a limited number
    of cell generations (usually 20-80
    population doublings) are known as
    finite cell lines.
    These cell lines exhibit the property of
    contact inhibition, density limitation
    and anchorage dependence.
    The growth rate is slow and doubling
    time is around 24-96 hours.
    Continuous cell lines 
    Cell lines transformed under laboratory
    conditions or in vitro culture conditions give
    rise to continuous cell lines.
    These cell lines show the property
    of ploidy  (aneupliody or heteroploidy),
    absence of contact inhibition and anchorage
    dependence.
    They grow either in a monolayer or in
    suspension (see below). The growth rate is
    rapid and doubling time can be 12-24
    hours. 
    Comparison of properties of finite and
    continuous cell lines:
    Basic cell culture
    techniques
     In the laboratory, cells are generally
    maintained in a suitable culture medium
    either solid or liquid, but in either case, one
    providing an environment suitable for their
    growth and multiplication.
     Furthermore, cultures of cells are normally
    supplied as pure cultures, containing only a
    single strain.
    This is in contrast to natural environments
    where a diversity of microorganisms is usually
    present.
    Techniques
    In order to maintain pure
    cultures, all glassware must be
    sterilized before use
    and aseptic
    techniques (sometimes
    known as sterile techniques)
    should be observed
    throughout, as described in the
    following notes.
    Basic cell culture
    techniques:
    Transfers of cells from one vessel to
    another should be carried out as
    rapidly as possible, though without
    unnecessary haste and without the
    need of a partner to hold anything.
    Plugs and caps must be held in the
    fingers when temporarily removed
    from culture vessels and must be
    set down on the bench.
    Basic cell culture
    techniques:
    The mouths of culture tubes or
    bottles should be flamed after
    removing plugs and caps and again
    before they are replaced.
    Work in a laminar flow or close to a
    Bunsen flame to ensure that
    airborne contaminants are carried
    upwards.
    Growing of cells (functions of
    media components)
    Cell Growth
    Lag Phase: 
    The time the cell
    population takes to
    recover from sub
    culture, attach to the
    culture vessel and
    spread.
    Initially growth is
    slow while the cells
    mobilize their stored
    food and their
    enzymes, but the
    Growing of cells
    Log/Exponential
    phase: 
    In this phase the cell
    number begins to
    increase exponentially
    provided that
    nutrients are in good
    supply, with cell
    numbers doubling as
    often as every ten
    minutes (although
    varies depending on
    cells).
    Growing of cells
    Plateau Phase: 
     During this phase,
    the growth rate
    slows or stops due to
    exhaustion of growth
    medium or
    confluency. As the
    nutrient supply is
    used up and as
    waste products
    accumulate, the rate
    of growth slows until
    the rate at which
    new cells are
    Factors affecting growth - Nutrients 

     Cells consist mostly of carbon, hydrogen,


    oxygen and nitrogen with smaller quantities of
    Sulphur and Phosphorus. These six basic
    nutrients are essential to growth.
     In addition, substances needed in much smaller,
    but still significant amounts are Potassium, Iron,
    Calcium and Magnesium (macronutrients).
     Needed in smaller quantities still are Copper,
    Zinc, Cobalt, Manganese and Molybdenum
    (micronutrients).
     Also required are a group of chemicals known
    as growth factors, which include vitamins,
    amino acids, purines and pyrimidines.
    Factors affecting growth –
    Temperature, pH
    Temperature – most cells grow
    most rapidly in the 20 – 45oC
    range.
    pH – Cells can survive a smaller
    range of pH than bacteria, being
    able to tolerate acidity as low as
    pH 6.0 and alkalinity pH 7.5.
    Factors affecting growth
    - Oxygen
     Most cells are aerobic and
    therefore need to be supplied
    with oxygen to grow (obligate
    aerobes). Some find oxygen
    toxic and grow badly in its
    presence (obligate
    anaerobes), while others grow
    better when oxygen is present,
    but continue to do so if its
    Factor affecting growth -
    Water
    All cells require water to grow,
    and in order to absorb it the
    water potential of the medium
    on which they exist must be
    higher (less negative than their
    cell contents.
    In these circumstances the
    medium dehydrates and
    thereby kills the
    Factor affecting growth -
    Light
     Some cells (plant)
    are photosynthetic and therefore
    require light.

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