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Thermo Scientific™ Restore™ Western Blot Stripping Buffer
Description
Thermo Scientific™ Restore Western Blot Stripping Buffer safely and effectively removes primary and secondary antibodies from nitrocellulose and PVDF membranes to allow chemiluminescent Western blots to be reprobed.
Performing gel electrophoresis and duplicate immunoblot assays to test new primary antibodies or antibody concentrations is time-consuming and expensive. Restore Western Blot Stripping Buffer eliminates this waste when detecting immunoblots with chemiluminescent Western blotting substrates. The reagent allows clean and efficient removal of primary and secondary antibodies from immunoblots without removing or damaging the immobilized antigen. This allows blots to be stripped and reprobed with confidence.
Highlights:
- Saves time – no need to re-run gels and blots
- Saves costly sample – re-probe the membrane using the same target sample
- Effective – formulation is more efficient at stripping antibodies than homemade buffers
- Gentle – does not damage the target antigen during stripping allowing efficient reprobing
- Odor-free – no mercaptans means no acrid odors
- Economical – less expensive than other commercial stripping buffers
Reuse a nitrocellulose or PVDF blot to detect a different target with a different primary antibody; Reprobe a blot to correct or optimize antibody concentrations that were ineffective the first time.
Specifications
Specifications
| Form | Liquid |
| Product Line | Restore |
| Quantity | 4 x 500 mL |
| Content And Storage | Storage: Upon receipt store at room temperature or 4°C. Product shipped at ambient temperature. |
Frequently Asked Questions (FAQs)
For nitrocellulose or PVDF membrane following Western blot detection using a chemiluminescent or fluorescent substrate system: Following transfer, air dry the membrane and place in an envelope, preferably on top of a supported surface to keep the membrane flat. The blot can be stored indefinitely at -80 degrees C. When ready to reprobe, prewet the PVDF blot with alcohol for a few seconds, followed by a few rinses with pure water to reduce the alcohol concentration. Then proceed as normal with blocking step.
FOR STRIPPING/REPROBING OF MEMBRANES:
Harsh protocol (see NOTE below for modifications)
1) Submerge the membrane in stripping buffer (100 mM BME, 2% SDS, 62.5 mM Tris-HCl, pH 6.7) and incubate at 50 degrees C for 30 min with occasional agitation. If more stringent conditions necessary, incubate at 70 degrees C.
2) Wash 2 x 10 min in TBS-T/PBS-T at room temperature.
3) Block the membrane by immersing in 5% blocking reagent TBS-T or PBS-T for 1 hr at room temperature.
4) Immunodetection
NOTE: Often you don't need such harsh conditions to remove antibodies from their proteins. The stringency of one or several of the variables can be decreased: lower the temperature, decrease the time, less BME, less SDS, etc. An especially mild but still often effective stripping protocol is lower pH incubation. Example: pH 2.0 Tris 50-100 mM, 30-60 min incubation (you may do two incubations if you wish). Then rinse and block as usual. If you do not wish to re-use the membrane immediately after stripping, you can store the membrane in plastic wrap (wet, you do not want it to dry out). Another simple, mild stripping buffer is 0.1 M glycine•HCl (pH 2.5-3.0), incubation 30 min to 2 hrs room temperature or 37 degrees C, depending on the antibody.
For Research Use Only. Not for use in diagnostic procedures.