We are trying to transfect BCPAP cells with a vector containing CMV-GFP- tumor suppressor gene but 95% of cells die less than 24 hours after transfection. We tried the following alternatives to solve the problem:

1-      Lipofectamine without any DNA ( cells all were completely healthy)

2-      We checked cells with mock (vector without tumor suppressor gene) but almost all cells were died

3-      In 96 wells plate we used 100 ng of DNA per well (according to protocol). it was diluted as following : 1/2, 1/4, 1/10 and 1/20 (cells were died at ½ , ¼ and at 1/10 ,1/20 dilutions, no significant GFP expression was observed albeit the cells were viable.

4-      Although we tried changing media of mentioned conditions after 6 and 8 hrs. but cells were yet died.

5-      For evaluating the influence of CMV, prostate specific vector with the same tumor suppressor gene with no protein expression in thyroid cells ,were used. However, we didn’t recognize any GFP expression but cells were completely died again.

DNA was also checked for probable contamination with endotoxins but nothing was detected.