BLS 201: CELL AND TISSUE CULTURE TECHNIQUES
SEMESTER III, END OF SEMESTER EXAMINATION, MARCH 2011
Question 1:
A 25 ml cell stock suspension was obtained after trypsinization of confluent Vero cells. 100 µl of the stock cell suspension was then added onto 900 µl of RPMI-1640. From the cell-RPMI-1640 mixture, 200 µl were added onto 400 µl of trypan blue and cells were then added onto a cell counting chamber. A total 300 cells (of which 50 stained blue with trypan blue) were seen in the five hemocytometer counting grids.
- Calculate the total number of cells left in the stock cell suspension (10 marks)
- Calculate the volume (in µl) of the stock cell suspension made after trypsinization required to make 10 ml of cell suspension at 1 × 105 cells/ml (10 marks)
Question 2:
Write short notes on the following:
- Cryopreservation of cells (4 marks)
- Cell counting (4 marks)
- Continuous cell lines (4 marks)
- Virus inoculation (4 marks)
- Colour development during IPMA (4 marks)
Question 3:
Provide the function(s) of the following equipments and reagents in cell culture;
| a) | Ice | (2 marks) |
| b) | Sucrose | (2 marks) |
| c) | Carbon dioxide (CO2) | (2 marks) |
| d) | Fluorescence microscope | (2 marks) |
| e) | Paraformaldehyde | (2 marks) |
| f) | FACS | (2 marks) |
| g) | PBS | (2 marks) |
| h) | Nystatin | (2 marks) |
| i) | Liquid nitrogen canister | (2 marks) |
| j) | Glycerine DABCO | (2 marks) |
Question 4:
Briefly explain how you would perform the following:
- Detection of viral infection in cultured cells (10 marks)
- Splitting and passaging confluent cells cultured in a cell culture flask onto a new cell culture flask (10 marks)
Question 5:
- Write short notes on virus titration by using plaque assay (10 marks)
- What is the virus titre of herpes simplex virus estimated by plaque assay in a 24- well plate performed by inoculating 100 µl of the virus at 1 X 107 dilution onto each well with Vero cells, given that the average number of plaque forming units (pfu) per well is 34? (10 marks)