BIOL114- Lecture 9 - Animal cell Biotechnology

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  • Created by: Katherine
  • Created on: 16-03-16 17:00
What are the uses and advantages of working with cultured cells over intact organisms?
More homogeneous than cells in tissues, Can control expermental conditions, can isolate single cells to grow into a colony of genetically homogeneous clone cells, can manipulate cells, can examine effects of treatment, no ethical issues.
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How do you generate a primary culture?
Primary cell cultures are established from animal tissues. Certain types of cells are easier to culture than others.
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What are the stages of tissue culture?
Generate a primary culture, grow to produce a primary cell line. Most cells undergo senescence, a few cells escape senescene andd form a cell line.
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What are the characteristics of transformed cell lines?
Immortal, anchorage independent, reduced requirement for growth factors, motile, no contact inhibition of growth, tumouigenic, less differentiated.
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What does it mean to be anchorage independent?
It does not require a surface to grow on
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What does it mean to be motile?
It can move around the culture dish
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What does it mean to be tumouigenic?
It will form tumours if injected into mice
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What does it mean to be less differentiated?
It loses specialised characteristics.
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Essentially, transformed cell behave in the same way as...
Cancer cells
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How do we grow cells in culture?
Requires rich growth medium containing esential amino acids, vitamins, salts, glucose. Sterile conditions. Generally need some form of serum (GF), Need to use orrect pH and osmolarity. Most grow only on special solid surfaces.
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Cultured cells can be used to generate what?
Proteins e.g. Antibody generation.
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Two different cells can be induced to fuse, therby creating a...
hybrid.
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Certain ... cells are used to produce...
Hybrid cells (hybridomas), monoclonal antibodies.
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What is a monoclonal antibody?
This is
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How can you make a monoclonal antibody?
Mouse is immunised with antigen of interest, once immune response has developed, the spleen is removed. Spleen cells are mixed with immortal cells from a mouse cell line (lymphoma). Some B cells fuse with lymphoma to create hybridoma.
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What happens to the cells that have no fused?
They cannot crown and are killed by a selective HAT medium.
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What happens to the hybridoma surviving cells?
They are separated out so that there is just one cells in each well of a culture dish and grow on to produce hundreds of cell lines. These lines are screened for the required antibodies.
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What are the uses of monoclonal antibodies?
Research (detect and measure specific proteins or other antigens), research (purify antigens by affinity chromatography), Diagnosis - detect markers of disease. Therapy e.g. Herceptin treatment of breast cancer.
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What was the first tissue engineered organ?
The trachea.
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What is a stem cell?
An undifferentiated cell. It is multipotent.
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What is a precursor cell?
It is a partially diffeentiated (unipotent) cell - it is not multipotent like the stem cell
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What happens to precursor cells?
They go on to become specialised.
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Where are stem cells found?
Either in the early human embryo at blastocyst stage or from adult bone marrow
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Where are embryonic stem cells derived from and what is their potency?
The inner cell mass - these are pluripotent
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Where are adult stem cells derived from and what is their potency?
Tissues (bone marrow), these give rise to a limited number of cell types - they are multipotent.
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What are the advantages to using embryonic stem cells? (ES)
They are pluripotent - can form any cell type
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What are the disadvantages to using Embroynic stem cells?
They are not genetically identical to patient, difficult to culture and there are ethical/moral issues associated with them.
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What are the advantages to using adult stem cells?
They are the patients own cells so are genetically identical. There are no ethical concerns.
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What are the disadvantages to using adult stem cells?
They can form a limited number of cell types. They are not available for all tissues.
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What is an ips cell?
An induced pluripotent cell
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How do you make an ips cell?
Introduce the four stem cell master regulator genes into a percursor cell, using the retroviral cloning vector. This switches the stem cell genes of the precursor cell on.
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How can ips cells be introduced to a patient?
Remove skin cells from patient, reprogram skin cells so they become induced pluripotent stem cells. Treat ips cells so that they differentiate into a specific cell type. eturn cells to patient, where they can repair damaged tissue.
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Are induced pluripotent stem cells the way forward? What are the benefits?
There are fewer ethical considerations, you can use a patients own cells, fully differenticated cells can be induced to become like ES cells.
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What are the risks associated with iPS cells?
They are technically challenging (cells are genetically engineered). There is a risk of cancer and oncogenes are induced.
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What is an alternative to ips cells?
Direct reprogramming - skin cells (fibroblasts) infected with viruses expressing neural precursos-specific transcrption factors.
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How do you generate a primary culture?

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Primary cell cultures are established from animal tissues. Certain types of cells are easier to culture than others.

Card 3

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What are the stages of tissue culture?

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Card 4

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What are the characteristics of transformed cell lines?

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What does it mean to be anchorage independent?

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