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Hello listers,
We have been working on refractive index matching following the
direction from Staudt, etc 2007.
[Reference] Thorsten Staudt, Marion C. Lang, Rebecca Medda, Johann
Engelhardt, and Stefan W. Hell. 2,2'-Thiodiethanol: A New Water Soluble
Mounting Medium for High Resolution Optical Microscopy. Microscopy
Research and Technique 70:1-9 (2007).
We prepared 10 %, 25 %, 50 %, and 97 % of TDE solution as stated in the
above reference. Linear relationship between the refractive index and
TDE concentration was confirmed by several measurements. However, we
have encountered two issues.
1. During the preparation of pure TDE (Thiodiethylene glycol from Fluka
88559) at pH 7.5 it was found out that the titration process is very
difficult. As stated in the above literature, we put 1 micro liter of
HCl (1.2 N). The pH was significantly changed to close to 2. Thus, we
dilute the HCl solution to 0.6 N. However, the changes of pH was also
significant. To compensate the pH level, 1 micro liter of NaOH (1 N) was
put into the very acidic solution. Then the pH went up to 9. From this
point, we prepared a very dilute HCl solution (0.015 N). Through series
of titration with 5 micro liter at each time the pH of the TDE solution
was adjusted to 7.42.
2. Sections of mouse brain with DNA stain or fluorescent Nissl were
prepared and kept in a refrigerator. After overnight storage, crystal
was formed in the slide. So we prepared another sample and kept in the
air to see whether temperature affects. Still, crystal was formed. Yet,
we don't know the reason for formation of crystals in the slides.
If you have any similar experience and overcame it, please let us know.
Thanks very much.
Best regards,
Ohkyung Kwon
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