Editing Transfection Reagent QC

1. Make a serial of dilutions in water, put concentrated transfection reagent back to 4 oC fridge.<br> 

{| cellspacing="1" cellpadding="1" border="1" style="width: 765px; height: 243px;"
|-
| Dilution<br> 
| Transfection Reagent<br> 
| Water<br> 
| Transfection Efficiency&nbsp; 12 hours <br>
| 
Transfection Efficiency 24 hours<br>

| 48 hours
|-
| 1:5<br> 
| 1 µl<br> 
| 4 µl<br> 
| <br>
| <br> 
| 
|-
| 1:10<br> 
| 1 µl<br> 
| 9 µl<br> 
| <br>
| <br> 
| 
|-
| 1:20<br> 
| 1 µl<br> 
| 19 µl<br> 
| <br>
| <br> 
| 
|-
| 1:30<br> 
| 1 µl<br> 
| 29 µl<br> 
| <br>
| <br> 
| 
|-
| 1:40<br> 
| 1 µl<br> 
| 39 µl<br> 
| <br>
| <br> 
| 
|-
| 1:50<br> 
| 1 µl<br> 
| 49 µl<br> 
| <br>
| <br> 
| 
|-
| 1:75<br> 
| 1 µl<br> 
| 74 µl<br> 
| <br>
| <br> 
| 
|-
| 1:100<br> 
| 1 µl<br> 
| 99 µl<br> 
| <br>
| <br> 
| 
|}

2. Add 3 µl diluted transfection reagent 50 µl DMEM (serum free).<br> 

3. Add 1 µg GFP or RFP&nbsp; plasmid DNA to 50 µl DMEM (serum free).<br> 

4. Mix both solutions, and incubate at RT for 15 ~ 20 min.<br> 

5. Add to one well 293T cells (12 well-plate, 60~90% confluency). <br> 

6. Check GFP or RFP expression 1, 2, 3 days after transfection under microscope.

7. Culture for 2 days, take a picture of each well under microscope.<br> 

8. Let cells to culture 2 more days, take a picture of plate using blue light transilluminator.<br> 

9. Check if cells have been contaminated by bacteria.<br> 

10. Make dilution of transfection reagent&nbsp; in 1 X PBS, filter through 0.22 µ filter in the biosafety hood.<br> 

11. Make 1 ml (for sale) and 100 µl (sample) aliquotes. <br> 

12. Record QC data on electronic notebook, upload images to notebook. 

13. Print labels, use date as lot number, put label on tubes. 

14. Store transfection reagent tubes at 4 <sup>o</sup>C. 

<br> 

[[Category:Protocols]]