Hoefer HB1000 - Instrukcja (Strona 28)

•
p19
Non radioactive probes
1. Wash blots in 2X SSC, 0.1% SDS for 3 minutes
at room temperature (repeat one).
2. Wash filter in 0.15X SSC, 0.1% SDS for
15 minutes at 50 °C (repeat once).
3. Store blots in 2X SSC at room temperature.
Radioactive probes
1. Remove blot from hybridization bottle and transfer
to Tupperware container.
2. Rinse briefly in 50 °C 0.1X SSC, 0.1% SDS.
3. Remove this solution and discard in radioactive
waste.
4. Wash blot three more times in the same solution.
5. After the final wash, dry the blot on filter paper
for 10 minutes. This is a good time to quickly
pass your hand held radioisotope reader (beta or
gamma counter) over your blot to get a general
idea as to the exposure time you will need for the
X-ray film. Hot blots can be placed against film for
20 minutes to 2 hours. Otherwise, exposure can
continue overnight (8 hours).
6. Tape the blot to a cardboard backing.
7. Cover with plastic wrap to prevent the blots from
sticking to the X-ray film.
8. Place the cardboard containing the blots into the
X-ray film folder.
9. In the darkroom, place a sheet of X-ray film over
the blot(s).
10. Place an intensifying screen on top of the film.
11. Close the film holder and clamp it.
12. Store at -70 °C. The low temperature reduces
light scattering and increases the length of
exposure time. Expose the blot for 20 minutes
to 24 hours.