North2South™ Hybridization Buffer

Yes. The most common procedure for stripping DNA blots is to wash the blot 3 x 5 minutes with boiling 0.1% SDS. An additional alkaline wash of 3 x 5 minutes with 0.4 N NaOH, 0.1% SDS may also work for DNA blots. The blots must be neutralized in TE buffer after this wash method. RNA blots are best washed with 0.1 % SDS, 3 x 5 minutes, at 75-85°C. Commercial stripping buffers are also available. Stripped blots should be kept wet until re-probed by storage in TE buffer at 4°C. Blots may be stripped and re-probed several times.

Yes. The hybridization buffers for the North2South Chemiluminescent Hybridization and Detection Kit contain single stranded nucleic acids used to block the membrane during the pre-hybridization and hybridization procedures. The addition of more single stranded DNA or RNA is not needed.

Yes. Certain components are offered separately: Cat. No. 17295, North2South Chemiluminescent Substrate for HRP, Cat. No. 37549, North2South Hybridization Buffer, Cat. No. 37555, North2South Hybridization Stringency Wash Buffer (2X).

Yes. A CCD Camera will work with the North2South Chemiluminescent Hybridization and Detection Kit (Cat. No. 17097). The results will depend on the quality of the chip in the camera. The peak light emission for chemiluminescence is at 425 nm. Traditional phosphorescence imagers do not work for chemiluminescence.

Yes. Standard colony lift protocols will work using a biotinylated probe. However, care should be taken to completely remove any residual bacterial debris or agarose from the filters, as this will cause high background signals. Non-specific speckling may be eliminated by centrifugation of the streptavidin-HRP conjugate in a bench-top microcentrifuge for several minutes at maximum speed before adding it to the hybridization buffer.