UVP HybriCycler - Manual (Page 14)

HC-3000 HybriCycler Hybridization Oven 14
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 piece of X-ray film over the blot(s).
10. Place an intensifying screen on top of the film.
11. Close the film folder 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.
Protocol 3: Chemiluminescence detection: UVP BioWest ™, HRP-
tagged, Alkaline phosphatase (AP) probes or antibody conjugates
Equipment
Clear plastic cling-wrap or Clear transparent sheet protector
UVP EpiChemi II™ Darkroom
Pipette
Cooled CCD camera
Reagents
UVP BioWest™ – For HRP Tagged Species
o BioWest Substrate Working Solution: Prepare by mixing equal
parts 1:1 BioWest Luminol/Enhancer Solution and BioWest Stable
Peroxide Solution. This working solution is stable for a minimum
of 24 hours at room temperature and may be stored in ambient
light and temperature.
BioWest Procedure
1. Immerse the blot in BioWest Substrate Working Solution for five
minutes. Completely immerse the blot in the substrate (0.125
mL/cm2).
2. Remove the blot from the substrate solution and place it in a sheet
protector.
3. Remove any bubbles between the blot and the surface of the
protector.
4. Place in an EpiChemi II Darkroom and with all internal lights off image
the blot.
Other Chemiluminescence
ECL tm (or other) detection reagent 1
ECL tm (or other if required) detection reagent 2