View source for Transduction of THP-1s

'''Transduction Reaction'''

* The volume of the reaction is based on the amount of virus available, the number of cells and the MOI being used   

*   Prepare the mixture as follows:
**  X uL of concentrated virus
**  X uL of SF Macrophage Media (chose this volume based on the vessel the cells will be grown in
** 1000X polybrene


'''Protocol'''

# Count cells, add cells to appropriate size tube, spin down at 400 x G for 5 minutes
# Re-suspend cells in the transduction reaction mixture
# Spin at 400 X G, 10 degrees for 1 hour 30 minutes
# Re-suspend the cells in the virus containing media and transfer to an appropriate size flask or plate, incubate overnight at 37 degrees overnight
# Transfer the contents of the flask to a new tube, spin down and replace the media with 500 uL of fresh complete RPMI
# Incubate plate for 48-72 hours at 37 °C


'''Notes'''

* Number of cells: Select number of cells that allows for an MOI of 300, some literature suggests MOI up to 500
* If making a cell line add selection media at the incubation period in step 6. I would also suggest comparing live/dead cell counts for both the control and the transduced cells before every media change to determine if the selection process is working