Fluorescein

1. Meet (or Re-Meet) Fluorescein
http://en.wikipedia.org/wiki/Fluorescein

2. “The fluorescence of this molecule is very high, and excitation
occurs at 494 nm and emission at 521.”
http://en.wikipedia.org/wiki/Fluorescein

3. Absorption and Emission of Light
From: Molecular Probes

4. http://www.invitrogen.com/site/us/en/home/References/Molecular-Probes-The-Handbook/pH-Indicators/Probes-Useful-at-Near-Neutral-pH.html#head1

5. Obtain fluorescein sample from TA; blank with appropriate buffer & re-blank
when you switch to a different pH. Determine appropriate dilution to use, and
scan from 200 – 600 nm. Move cursors to obtain absorbance values at a particular
lambda max.
Analyze your data using
(1) Nonlinear fitting
(2) Linear fitting
(3) 2nd Derivative fitting
To speed things up, we will read at 240 & 490 nm
rather than scanning from 200 – 600 nm.