Never tried this before, but I'm sure someone out there has done something
similar. We could use some advice to prevent us from wasting our $$$.
We are using a substrate which contains a fluorogen to detect the presence
of our favorite enzyme. As a result of the enzymatic reaction,
p-nitroaniline is produced and can be detected via fluorescence. While we
have an automated plate-reader that can do fluorescence readings, we need
to buy the appropriate filter(s) (for the platereader) to get the proper
wavelengths to work with p-nitroaniline.
The literature is a bit confusing on this point. Some sources cite an
excitation wavelength of 340 nm; others 355 nm; and one even said 405 nm
(although this last one might be to detect p-nitroaniline via simple
If anyone out there is well-versed in fluorescent detection of
p-nitroaniline, can you suggest an optimal excitation wavelength (and the
cognate emission wavelength)?
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