Animal cell technology

• What is homogenous


  one layer thick
• why use cell culture

  can control

  can isolate single cells which can grow into a colony of genetically homogenous clone cells

  can manipulate

  can examine effects of treatments of intracellular events more easily

  cheaper

  no ethical issues
• How to grow cell in culture?

  isolate cells

  1. use a growth media with amino acids, vitamins, salts, glucose
  2. They are typically included with serum (fetal bovine serum)
  3. NEED sterile conditions (double time human 24-30 hrs)- so no out competition from bacteria
  4. Need correct pH and osmolarity - so use buffers (phosphate-buffered saline)
  5. Most media has an indicator (usually indicating pH has changed)
  6. grow on special solid surfaces
• What occurs when cells use up all growth materia

  they release co2 and other things that cause a toxic external environment
• stages of tissue culture:

  1. generate a primary culture
  2. growth cells to produce a primary cell line
  3. senescene
  4. few cells form a cell line
• Which cells are appropriate for cell culture

  hardy skin cell are a good example
• how to establish cells for a primary culture

  Extract cells from tissue using trypsin (protease)
• what is contact inhibition

  when cell are in contact with out cells or surface of plate they will stop growing
• How to make a cell culture last longer

  In order for them to keep growing, put more trypsin into cell plate, which deattaches them from dish and together,

  Put in to fresh media
• Why do some cells escape senescene

  mutation
• What are the cell called that escape senescene

  cell line (cancerous)
• Positive characteristic of cell line

  immortal
• who is Henrietta Lacks

  Had cells stolen from her carcinoma of uterine cervix, which is used for cell development
• negative characteristics of cell lines

  Anchorage independent – do not require a surface to grow on

  Reduced requirement for growth factors

  Motile – move around culture dish

  No contact inhibition of growth – cells grow on top of each other

  Tumourigenic – will form tumours if injected into mice

  Less differentiated – lose specialised characteristics
• What can you do with cell lines

  protein production

  tissue engineering
• Describe cell line making proteins

  antibody production

  (polyclonal and monoclonal)
• How to generate a monoclonal antibody

  Form a hybrid cell (two cells fused together) = hybridomas

  Mouse tumour cells are cultured fused with a mouse spleen cells
• use of monoclonal antibodies

  Research (for detection or purify antigens by affinity chromatography)

  diagnosis

  therapy (herceptin)
• sources of stem cells

  embryonic stem cell

  adult stem cell
• Advantages and disadvantage of embryonic stem cells

  •        Advantages: Pluripotent – can form any cell type in the human body

  •        Disadvantages: Not genetically identical to patient, Difficult to culture, Ethical/moral difficulties
• Advantages and disadvantages of adult stem cells

  •        Advantage: Patient’s own cells – genetically identical

  •        No ethical concerns

  Disadvantages:

  •        Multipotent: Form limited number of cell types
• master regulator cells:

  Oct3/4

  Sox 2

  Klf4

  c-Myc
• How to produce induced pluripotent stem cells:

  remove skin cells

  reprogram

  treat cells so they differentiate

  return cells to patient where they can repair damage tissue
• Direct reprogramming:

  Somatic cell to become another cell type directly
• How to use direct reprogramming

  Transgene overexpression using viruses

  changing endogenous gene expressing using CRISPR /cas 9 or pharmacological agents