Cell and Tissue Journal

Abstract Aim: Different methods have been used for introducing DNA into mammalian cells. Calcium phosphate-based transfection protocols are inexpensive and easy to perform, however, the transfection efficiency in some cases is not satisfactory. In this study we try to increase the efficiency of calcium phosphate-mediated cell transfection.   Material and Methods: 2.0×10⁵ HEK293T cells were plated in each well of a 6-well plate containing a 10% poly L lysine-coated coverslip and grown to 60% confluency. The cells were then transfected with a plasmid encoding GFP (Green Fluorescence Protein), using a standard calcium phosphate protocol in the presence or absence of chloroquine phosphate. GFP protein expression was observed by immunofluorescence microscopy and quantified with the help of Image J software.  Results: We have shown that chloroquine phosphate can highly improve the efficiency of calcium phosphate-dependent transfection in the HEK293T cells, leading to an approximate 8-fold increase in the number of transfected cells. Chloroquine phosphate was not toxic to the cells, however, its positive effect on transfection efficiency was abolished in the presence of glycerol or DMSO (Dimethyl sulphoxide). Conclusion: we recommend chloroquine phosphate for increasing the efficiency of transfection by the inexpensive method of calcium phosphate. In addition, we recommend that chloroquine phosphate to be used alone and not along with other inducers of cell transfection.