Procedure for transfecting L929 cells

L929 is a murine aneuploid fibrosarcoma cell line which is often used to assay TNF-alpha and TNF-beta. Treatment with TNF initiates apoptosis and subsequent cell death, therefore L929 cell is often used for immunological assays. However L929 cell is hard to transfect, especially resistant to liposome based transfection method. We have been using GenJet Ver. II and PolyJet to transfect L929 cell and got up to 75% efficiency. The brief procedures are described below for transfecting L929 cells with GenJet and PolyJet.
1. Grow L929 cell to ~80% confluency at the day of transfection. L929 cell must be healthy and less than 9 passages for maximum efficiency.
2. For 6-well plate, dilute 1.0
µg of DNA and 3.0 µl of GenJet Ver. II or PolyJet reagents per well with serum free DMEM respectively. Add diluted reagent to diluted DNA and let transfection complex formed at RT for 15 minutes.  For other format of cell culture formats, scale down or up per the surface area of culture dish.
3. Add transfection complex to L929 cell directly. Transfection is conducted in presence of serum/antibiotics with transfection volume of 1.0 ml per well of 6-well plate.
4. Check transgene expression 24~48 hours post transfection.


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