How do you perform lentiviral transduction?

c. Transduction

Thaw the lentivirus on ice. Mix 8 µl Polybrene (1 mg/ml aliquot) with 957 µl culture.
The next day, exchange Lentivirus/Polybrene mixture by fresh culture medium. Incubate cells at standard cell culture conditions.
concentrations range from 0.1-10 μg/ml. Replace the culture medium 48-72 hours.

How do you calculate Tu mL?

Method 1: TU/mL = (Number of cells transduced x Percent fluorescent x Dilution Factor)/(Transduction Volume in mL)
Method 2: TU/mL = (Number of cells transduced x Percent fluorescent)/(Virus volume in mL)

How long is incubation for lentivirus?

9- Do not forget the incubation time All lentiviral vectors present in the transduction mix need at least 5 hours to penetrate the cells of interest. Based on the experiment, the transduction can be left from 5 hours to an overnight incubation.

How is transduction efficiency calculated?

To determine transduction efficiencies, a total of 5 × 104 cells (adherent cells) or 1 × 105 cells (suspension cells) per well in 24-well plates were inoculated with serial dilutions of the virus supernatants overnight.

What is the difference between transfection and transduction?

Transfection is the process of introducing nucleic acids into cells by non-viral methods. Transduction is the process whereby foreign DNA is introduced into another cell via a viral vector.

Do you need to snap freeze lentivirus?

You can freeze lentiviral vectors, as straight supernatant from producer cells or after any method of concentration. Freeze rapidly (some recommend snap freezing in liquid N2 or dry ice / ethanol baths) and store at -80C.

What is the difference between transduction and transfection?

What is transduction efficiency?

For lentiviral constructs with a fluorescent marker or antibiotic resistance marker, transduction efficiency (i.e., % infected cells) can be determined from the fraction of fluorescent or antibiotic resistant cells in the population.