Question

This is the first-time you are doing tissue culture. You professor asked you to mix together a new bottle of DMEM with all the media components that she left for you in the tissue culture hood. You also received a brand-new bottle of Trypsin/EDTA, a new bottle of PBS and a tissue culture plate coated with collagen. Your professor then gave you a dish of adherent epithelial cells to passage. After following the passing cells protocol, you looked at your epithelial cells under the microscope 4 hours later and to your surprise not one cell was attached to the dish! This was surprising since cells usually attach to dishes within 2 hours.

A)What kind of microscope did you use view your cells in the culture dish?

B) Why does coating cell culture dishes with collagen allow cells to attach to the dish? (2 sentences max)

C) After discussing your results with your professor, she looked into the tissue culture hood and found a tube that contained one essential component you should have added to your DMEM media. What component did you forget to add and why did this prevent your cells from attaching to the dish? (3 sentences max)

Answer 1

Answer :

A)

Live cell tissue culture microscope.

Other microscopes give better results only after fixation and staining. In LCTC microscope, culture cells can be viewed in live form.

B)

Collagen type II and IV are used. This helps cell proliferation and adhesion. Adhesion occurs by interaction between cell adhesion molecules and transmembrane proteins on cell surface. Cell surface collagen receptors bind to collagen. Cellular fibronectin membrane proteins mediate attachment of cells to collagen substrates.

C)

Addition of calcium chloride dehydrate helps in cell adhesion. Calcium facilitates attachment of cells to substrate and one another. It modulates function of cadherin, selectin and integrin.

Collagen coated in dishes help the cell receptors to attach to it. Alfa poly L lysine also helps to adhere to the plate if not coated by collagen.