Hello, all!
I have calculated the percent of tranduced cells (%) and virus titer (TU/mL). I computed them after checking antibiotic selection process, which is a kind of functional titer, for 2 weeks. I will describe the calculation process. Additionally, I used a cell counter in order to calculate the percent of tranduced cells (%) and virus titer (TU/mL).
1. % of tranduced cells calculation
- Formula (%) = [{(antibiotic selected cells) - (cells in media only)} / {(non-antibiotic selected cells) - (cells in media only)}] x 100
- antibiotic selected cells : Antibiotic-resistant lentivirus and antibiotic were placed into target mammalian cells. After 2 weeks, the viability was 1.4 x 106 cells/mL.
- non-antibiotic selected cells : only antibiotic were placed into target mammalian cells, which do not have the resistance of antibiotic. After 2 weeks, the viability was 1.0 x 105 cells/mL.
- cells in media only : none of lentivirus and antibiotic were into the target mammalian cells. After 2 weeks, the viability was 1.5 x 107 cells/mL.
- Conclusion : [{(1.4 x 106 cells/mL) - (1.5 x 107 cells/mL)} / {(1.0 x 105 cells/mL) - (1.5 x 107 cells/mL)}] x 100 ≒ 91.3%
Thus, 91.3% of the target mammalian cells were transduced by antibiotic-resistant lentivirus.
2. virus titer calculation
First of all, I tried to use the formula, "TU/mL = (# of cells on transduction) x (MOI / mL of lentivirus stock)", but I have no idea what the MOI was. Therefore, I applied this formula, "TU/mL = # of colonies/mL x dilution."
- Formula (TU/mL) = # of colonies/mL x dilution
- # of colonies/mL : when I tried to do count the antibiotic selected cells in 1 mL media, the result was 1.4 x 106 cells/mL.
- dilution factor : I have grown the target mammalian cells in the 1 mL of complete media and the 1 mL of lentivirus stock, which is from 293FT cells.
- Conclusion : (1.4 x 106 cells/mL) x 2 = 2.8 x 106 TU/mL
Thus, the number of virus in the lentivirus stock is 2.8 x 106 TU/mL.
If I did something wrong on the calculation process, please tell me what they are.
I need your help, all.
Also, the reason why I did not use the flow cytometry was because no flow cytometry is set in my lab; consequently, I should try to do antibiotic selection process.
Please teach me, and help me.
Thank you so much for reading my questions.
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