Lentivirus Titration by 293T infection

Split cells

remove DMEM(+) medium (with 10% serum and PS) from T25 flask;

Add 5 ml PBS to the flask to wash the cells;

remove PBS;

Add 0.5 ml 1X trypsin;

Keep at room temperature until the cells are loose (1~2 min)

Add 5 ml DMEM(+) to the flask, use pipette to disperse cells by pipetting liquid up and down.

Add 10 ml DMEM(+);

Add 15 ul 1000X Polybrene

Transfer cells to 12 well plate, 1 ml/well, keep the last 3 ml, add 1 ml DMEM and continue to culture.

Infection

Add 1 ul, 2.5 ul, 5 ul or 10 ul lentivirus prep to one well.

Grow at 37oC in CO2 incubator for 2 days.

Take a picture of the cells under fluorescence microscope (to see how many cells are fluorescent).

Estimate infection rate.