Switch From a Traditional Hybridization Buffer to ULTRAhyb
ULTRAhyb® Ultrasensitive Hybridization Buffer can increase the sensitivity of a blot hybridization up to 100-fold.
If Using a Traditional Hybridization Buffer, Switch From DNA to RNA Probes
RNA probes are more sensitive than DNA probes when using traditional hybridization buffer. ULTRAhyb® Ultrasensitive Hybridization Buffer, however, substantially increases the sensitivity of DNA probes making them as sensitive as RNA probes.
Use Downward Alkaline Capillary Transfer Instead of Neutral Capillary Transfer
Alkaline transfer nicks the RNA, increasing sensitivity by more efficiently moving RNA, especially larger transcripts, from the gel to the membrane. Downward transfer does not crush the gel and makes use of gravity to speed up the process. (Ambion’s NorthernMax® Kits include reagents for alkaline transfer.)
Use an Optimal Hybridization Temperature
Temperatures of 42°C for DNA probes and 68°C for RNA probes usually give excellent results in 50% formamide-based hybridization buffers. Hybridization conditions for probes that have an unusually high GC or AT content or probes with a high degree of mismatch with the target should be determined empirically.
For Research Use Only. Not intended for human or animal therapeutic or diagnostic use.
The trademarks mentioned herein are the property of Life Technologies Corporation and/or its affiliate(s) or their respective owners.