I would like to perform 3-color immunofluorescence imaging on three proteins in cells. One of them is EGFP-tagged. The red color dyes (Texas Red or Alexa Flour 568) worked very well. However, the blue dyes (Alexa Flour 350 or 405) worked poorly. Any recommendation for another color dye?
I'd recommend using a Far-red, such as Alexa Fluor 647. They may not be able to be seen by eye, but they tend to be very bright and more difficult to photobleach while imaging. The actual color doesn't matter, because you can pseudo-color the images to complement each other.
the marine blue from life technologic (Catalog number: M-10165) works very well for me with the two dyes you have mentioned
I'd recommend using a Far-red, such as Alexa Fluor 647. They may not be able to be seen by eye, but they tend to be very bright and more difficult to photobleach while imaging. The actual color doesn't matter, because you can pseudo-color the images to complement each other.
If you do not have a 647 nm filter but a Dapi filter you can also use AMCA conjugated secondary antobodies or streptavidin and of cause CY2 or Alexa 488 (green coulor).
Good luck!
Thank you very much for all the useful answers. The Cy5-conjugated secondary antibody worked very well.
- Badges
- Science topic
- Similar topics
- Chemistry
- Biochemistry
- Biochemical Techniques
- Blotting Techniques
- Immunoblotting
- Dyes