Hello, I'm trying to visualize eGFP mice which have uptimum excitation wavelength at 488nm. It seems that there is no commercial flashlight or lamp that output this or similar wavelength. The usual hand-held UV light is ~360nm which doesn't work at all. The closest ones are 460nm and 470nm. Does anyone have the experience in eGFP mice visualization? Does 460nm work for eGFP? Thanks!
Hi Yue!
If You need just to visualize the whole GFP mouse, you can look at them in a dark environment using a blue led lamp (465-470 nm) and a green acetate filter. Use a wt as control. I bought my 3w lamp on ebay and the filter was a sample from http://leefilters.com/ (122 fern green). A filter for SyBr green gel imaging works as well.
Just to have an idea of the result, look at the attached pictures .
Good Luck
Franco
I visualized C57BL6/GFP mice with 360nm lamp (It should be totally dark all around) and I could see their ears and tail fluoresce in green color.
For more advanced uses you can check out IVIS whole animal imaging systems or other similar devices. (These imagers employ CCD cameras and detect low levels of photon emission). see link.
good luck
http://www.perkinelmer.com/catalog/product/id/ivisspe
Thanks Ehud!. I've tried 360nm in a dark room but it didn't work at all, while the mice glowed under 488nm in a gel imaging system in my lab. I'm trying to find a portable light source so I can take it to mouse house, because It's violation of the rules at my university to take alive mouse out, check them and put them back.
Dear yue,
I also used something like this apparatus (see link), it's portable.
I presume your problem is more related to the intensity of your lump or your GFP expression levels and not wave length.
If you just need an indication of positive GFP expressing mice (and all else fails :-() you can also do RTPCR from your mice with any tissue sample
Hi Yue!
If You need just to visualize the whole GFP mouse, you can look at them in a dark environment using a blue led lamp (465-470 nm) and a green acetate filter. Use a wt as control. I bought my 3w lamp on ebay and the filter was a sample from http://leefilters.com/ (122 fern green). A filter for SyBr green gel imaging works as well.
Just to have an idea of the result, look at the attached pictures .
Good Luck
Franco
Hi Yue Lu, I'm presuming you do not have access to a small animal imaging device (such as the IVIS) in your animal facility, as that would be the ideal system for imaging your animals. If you want to just look at the GFP expression the following system works pretty well:
The dark reader system - http://www.clarechemical.com/lamps.htm
It uses a Blue light that excites GFP pretty well & it has a orange filter that comes with it. The combination works pretty well - also I've taken pictures with a digital camera by putting the filter infront of the lense of the camera - and it comes out OK - not publication quality - but good enough for lab presentations.
The following one I have not used but BioRad claims it works for GFP:
http://www.bio-rad.com/en-us/product/uv-lights
http://www.clarechemical.com/lamps.htm
Hello Yue Lue,
I second Wasim H Chowdhurys recommendation of a blue light with an orange filter in front. I've been using a BlueStar flashlight from Nightsea to visualize fluorescent mice (C57BL/6-Tg(CAG-EGFP)1Osb/J) directly in the cage.
Good luck!
http://www.nightsea.com/products/bluestar-flashlight-packages/
Hi Dylan.
I never tried YFP, but a similar arrangement may work for RFP. Everything depends on the concentration (expression level) of the protein and the intensity of the excitation. RFP may be excited with green or blue leds and looked through an orange/red filter. The blue light gives a weaker excitation but a stronger contrast when observed with the red filter. The attached image is quite ugly, but shows both the purified proteins seen under these conditions.
All the best