I am trying to optimize how much amount of the BLOCK-iT™ Alexa Fluor® Red Fluorescent Control (Invitrogen, Catalog number: 14750100) I should use in my experiments. I have followed the reverse transfection protocol using Lipofectamine RNAiMAX in 6-well plates. After 34 hours , I sucked out the medium and added PBS, and then went onto image each well using fluorescence microscope.
1. I used Green filter for checking the fluorescence. Is this the right filter that I should use? Please suggest.
2. How can I say the efficiency of my transfection in terms of % from fluorescence microscopic examination? Should I count the cells in a haemocytometer? Right now, I have saved images in UV mode,fluorescence mode and the merged mode. From these images, is there any way to know the transfection efficiency?
Thanks in advance
Edited by Neanderthal, 19 October 2012 - 10:08 AM.