Lecture 3 Flashcards
What are the 4 basic things that must be controlled to grow cells in culture?
O2, temp, humidity, pH.
what can maintain the environment for cells?
Incubator.
what is the optimum pH for mammalian cells
7-7.4
Under what pH is mammalian cell growth inhibited?
6.8
What can regulate the pH of a medium?
Buffering systems, e.g. sodium bicarbonate-CO2. (bicarbonate added to medium, CO2 pumped in from gas tank when needed).
What is osmolality?
Concentration of a solute in a solution measured by mass.
Does osmolality affect cell growth and function?
Yes. Can be determined by components of growth medium.
What is Mr Frosty filled with before cryovials full of substrate are added?
Isopropanol.
Does the pH of the medium increase of decrease over time with mammalian cells? why?
Decrease, because of metabolic products, e.g. lactic acid.
What indicator can be used to determine pH of growth medium? What is the colour change on becoming too acidic?
phenol red.
Turns yellow.
Are established cells often aneuploid?
Yes.
Do established cell lines have normal doubling times?
No, often shortened.
What are 2 main ingredients in medium?
Glucose and glutamine.
Does CO2 increase or lower pH?
Lowers.
What is transformation?
Genetic modification allowing long term propagation in culture, causing a phenotypic modification.
What 4 does the extracellular matrix do?
Support, segregate, aid communication, act as a depot.
Does the matrix change over time? When? What enables this?
Yes, during growth and development (by use of proteinases and matrix metalloproteinases), and wound healing (fibroblasts).
Components of ECM?
Proteoglycans (e.g. chondroitin).
Polysaccharides (e.g. hyularonic acid).
Fibres (e.g. collagen, elastin).
Fibronectin, laminin.
Types of cell adhesion
Focal adhesion (ECM interacting w/ actin). hemidesmosome-ECM interactions. integrin.
Do cells bind to each other, or to filaments?
Both.
Do fibroblasts interact with the matrix?
Yes.
what happens to signals from the extracellular matrix when they bind to a receptor on a cell?
Signal transduction pathways -> cytoskeleton -> EITHER
Organisation and movement of molecules.
OR
Cell shape movement and contraction.
2 reasons to do adhesion assays?
Understand how tissue structures form.
Understand how tumours bind during tumour metastasis.
Basic mechanism of adhesion assay?
Coat microplates with matrix of choice.
Cells stained with colorimetric/fluorescent dye, usually calcein AM.
Cells added to micro plates, left, then washed to remove cells that have not adhered.
Fluorescence/colour then measured.
