Frequently Asked Questions
Q. What should be the maximum splitting ratio for any culture? Can it be more than 1:10? Find your answers here!
There is no thumb rule for the splitting ratio. It really depends upon the cell lines, you are working with. So, if you are using any normal, diploid human cells they may express a limited life span; so in that case, it should be noted that splitting at higher ratios may exhaust the available population doubling leading to rapid senescence.
On the other hand, some immortal cell lines like stem cells can be split at a higher ration. It is always advisable to prepare a standard growth curve with definite seeding density for each cell lines.
Q. How can you achieve an optimum growth environment? Name any two factors affecting the same?
Some of the basic components that are very crucial for the optimum growth potential of cells are being listed herewith:
- Amino Acids: The most wanted raw material promoting protein synthesis and cellular replication.
- Monosaccharide: Hexose formed through aerobic as well as anaerobic glycolysis is the primary source of energy in cultured cell lines.
- Inorganic ions and trace elements: The optimum growth of cells need some trace elements like molybdenum, vanadium, zinc, etc.
- Osmotic pressure: Cells need isotonic environment along with the ideal osmotic pressure i.e. about 290 mOsm/kg.
- pH: With the rapid fluctuation of pH more than 7.4 or lesser than 7.2 can be harmful to the cells.
Q. Is there any difference between cell lines, cell strains and cell types?
As per the nomenclature, the cell lines can be defined as the continuous propagation of cells that have undergone genetic transformation with infinite growth potential. A cell strain is a selective subpopulation isolated from a cell line isolated through single cell clones. They may have undergone an additional genetic transformation. Whereas, the term cell type can be cells with a common phenotype, say for example keratinocytes, melanocytes, etc.
Q. What is the best method to freeze primary cells, without much of a loss?
The best method to cryopreserve proliferating cultures can be with the use of a trademark product, the cryo-store; obtained from the stem cell technologies. The cryopreservation medium needs to be thawed at a temperature of 370c and should be lowered to 4 0c before being used. For better results, it is always suggested to use control rate freezers.
Q. How can I maintain good cell yield/viability during revival of cryopreserved cells?
Always remember slowly freezing and rapid thawing is the key for successful revival of cryopreserved cells. Post removals from liquid nitrogen immediately thaw the cryovial of cells in a 37°C water bath. Remove the cryovials from water bath when there some ice crystals are still remaining in the vial. Usually this entire procedure of thawing takes about 1-2 minutes.
Make sure to sufficiently dilute the DMSO after thawing cells before centrifugation (only at very low speed). As far as possible avoid centrifugation post revival.
Q. Can I use the growth medium available in my lab for cell growth?
Once you receive the cells please go through the safety data sheet for procedural details. We also suggest which medium to be used / cells grow better. We recommend same medium for growth of cells in your lab.
Q. What percentage of primary cell viability can be expected in Kosheeka’s vials?
Cell viability will be greater than 90% if you are choosing Kosheeka for your primary cell culture.
Q. What donor information can be obtained against a specific vial lot of primary cells?
Complete records for all species-specific and tissue-specific primary cells and stem cells are maintained with Kosheeka. Please contact us at firstname.lastname@example.org for any enquiries.
Q. Is it advisable to refreeze Kosheeka’s primary cells?
Refreezing the first stock will help you start from lower passage but frequent refreezing and thawing should be avoided at all costs to maintain good cell viability.
Q. How can I calculate how many cells should I seed for my cell culture experiment ?
The recommended seeding density for various flasks is as below.
|Flask||Area (mm2)||Cells seeding density|