Paul.xu at 16:20, 24 October 2011

2011-10-24T16:20:49Z

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

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

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

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

3. Add 1 µg GFP or RFP  plasmid DNA to 50 µl DMEM (serum free). 

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

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

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. 

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

9. Check if cells have been contaminated by bacteria. 

10. Make dilution of transfection reagent  in 1 X PBS, filter through 0.22 µ filter in the biosafety hood. 

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

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 oC.

 

[[Category:Protocols]]

Transfection Reagent QC - Revision history

Paul.xu at 16:20, 24 October 2011