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Restore™ Western Blot Stripping Buffer, Trial Size (Thermo Scientific™)

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.

Features of Restore Western Blot Stripping Buffer:

• 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

Product Details
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. Restore Stripping Buffer provides clean and efficient removal of primary and secondary antibodies from immunoblots without removing or damaging the immobilized antigen allowing blots to be stripped and reprobed with confidence.

Chemiluminescent Western blot detection with reagents such as Thermo Scientific SuperSignal Substrates for horseradish peroxidase is one of the most common and sensitive methods in use today. Because these substrates do not precipitate and bind to membrane surfaces, Western blots detected by chemiluminescence can be stripped with reagents that remove affinity-bound primary and secondary antibodies. To be effective, a stripping buffer must be strong enough to disassociate bound antibodies but gentle enough to leave the transferred target proteins intact on the nitrocellulose or PVDF membrane. Restore Western Blot Stripping Buffer has these characteristics.

By stripping and reprobing, there is no need to waste rare or costly samples by running multiple gels in order to probe for different targets. A single membrane from one gel can be stripped with Restore Western Blot Stripping Buffer to remove the primary antibodies. Stripping the blot takes only 15 to 30 minutes, depending on the affinity of the primary antibody. After stripping, block and reprobe with a new primary antibody. Alternatively, a blot can be stripped and reprobed with adjusted antibody concentrations to optimize conditions after obtaining initially poor results.

Applications
• 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

Protocol Summary
• Wash blot to remove chemiluminescent substrate.
• Incubate blot in Restore Western Blot Stripping Buffer for 5 to 15 minutes at 37°C (room temperature is sufficient for some antibodies).
• Remove blot and wash in Wash Buffer (TBS-T or PBS-T).
• Test for sufficient removal of antibodies.
• Perform next immunoblot experiment.

Restore™ Western Blot Stripping Buffer (Thermo Scientific™)

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.

Features of Restore Western Blot Stripping Buffer:

• 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

Product Details
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. Restore Stripping Buffer provides clean and efficient removal of primary and secondary antibodies from immunoblots without removing or damaging the immobilized antigen allowing blots to be stripped and reprobed with confidence.

Chemiluminescent Western blot detection with reagents such as Thermo Scientific SuperSignal Substrates for horseradish peroxidase is one of the most common and sensitive methods in use today. Because these substrates do not precipitate and bind to membrane surfaces, Western blots detected by chemiluminescence can be stripped with reagents that remove affinity-bound primary and secondary antibodies. To be effective, a stripping buffer must be strong enough to disassociate bound antibodies but gentle enough to leave the transferred target proteins intact on the nitrocellulose or PVDF membrane. Restore Western Blot Stripping Buffer has these characteristics.

By stripping and reprobing, there is no need to waste rare or costly samples by running multiple gels in order to probe for different targets. A single membrane from one gel can be stripped with Restore Western Blot Stripping Buffer to remove the primary antibodies. Stripping the blot takes only 15 to 30 minutes, depending on the affinity of the primary antibody. After stripping, block and reprobe with a new primary antibody. Alternatively, a blot can be stripped and reprobed with adjusted antibody concentrations to optimize conditions after obtaining initially poor results.

Applications
• 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

Protocol Summary
• Wash blot to remove chemiluminescent substrate.
• Incubate blot in Restore Western Blot Stripping Buffer for 5 to 15 minutes at 37°C (room temperature is sufficient for some antibodies).
• Remove blot and wash in Wash Buffer (TBS-T or PBS-T).
• Test for sufficient removal of antibodies.
• Perform next immunoblot experiment.

SuperSignal™ Western Blot Enhancer (Thermo Scientific™)

Thermo Scientific SuperSignal Western Blot Enhancer contains a membrane treatment reagent and a primary antibody diluent that increase both signal intensity and sensitivity 3- to 10-fold compared to a detection performed without it.

Features of SuperSignal Western Blot Enhancer:

Increase sensitivity—achieve 3- to 10-fold increase in signal intensity and sensitivity
Improve specificity—improves signal-to-noise ratio for poor quality and low affinity antibodies
Better clarity—reduces background for cleaner Western blots
Membrane compatibility—provides effective signal enhancement with PVDF and nitrocellulose membranes
Substrate compatibility—validated for use with chromogenic, chemiluminescent and fluorescent detection methods

When a protein or antigen is difficult to detect because of low abundance or poor immunoreactivity, use of SuperSignal Western Blot Enhancer can significantly reduce background and enhance detection of low-abundance and weakly immunoreactive antigens.

Power Blotter 1-Step™ Transfer Buffer (5X) (Invitrogen™)

Invitrogen Power Blotter 1-Step Transfer Buffer is designed for rapid semi-dry transfer of proteins from polyacrylamide gels (SDS-PAGE) to nitrocellulose or PVDF membranes using the Power Blotter System. Transfer is compatible with commonly used detection methods such as staining, chemiluminescence, or fluorescence.

Features of Power Blotter 1-Step Transfer Buffer:
Fast—high ionic strength formulation allows for 5- to 12-minute protein transfer when used with compatible fast semi-dry blotting systems
Optimized—designed for seamless and reliable performance with the Power Blotter System
Compatible—effective with other semi-dry transfer devices equipped with a suitable high-current power supply
Economical—no need for special or costly consumables that are required by other fast semi-dry transfer devices

Learn more about the semi-dry transfer device: Power Blotter System

When used with the Power Blotter System, this buffer provides effective gel-to-membrane transfer of proteins in 5 to 12 minutes with efficiency that is similar to conventional western blotting techniques. Fast blotting methods require a high-current power supply, such as the Power Blotter System, and an optimized high ionic strength transfer buffer, such as Power Blotter 1-Step Transfer Buffer.

By increasing the current, excellent transfer efficiency can be achieved in much shorter time compared to conventional methods. Amperage is held at a constant rate based on the surface area of the transfer stack(s) (~22–23 mA/cm2) and voltage is limited to 25 V.

Power Blotter 1-Step Transfer Buffer is also compatible with other protein semi-dry transfer devices, when they are paired with a suitable high-current power supply. Such devices provide constant high current (1.3 to 5.0 amps) to rapidly transfer proteins via the high ionic strength conditions supplied by the transfer buffer.

Related products
Power Blotter Pre-cut Membranes and Filters, nitrocellulose, regular size
Power Blotter Pre-cut Membranes and Filters, nitrocellulose, mini
Power Blotter Pre-cut Membranes and Filters, PVDF, regular size
Power Blotter Pre-cut Membranes and Filters, PVDF, mini

Restore™ Fluorescent Western Blot Stripping Buffer (Thermo Scientific™)

The Thermo Scientific Restore Fluorescent Western Blot Stripping Buffer is a gentle and highly effective reagent for quickly removing primary and near-infrared (IR) dye-labeled secondary antibodies from western blots.

Features of Restore Fluorescent Western Blot Stripping Buffer:

Fast—strip blots in only 15 minutes at room temperature
Saves time—no need to run new gels and prepare a new blot
Conserve samples—reprobe the same PVDF membrane for multiple targets
Economical—less expensive than other commercially available stripping buffers
Efficient—effectively strips blots the first time

Restore Fluorescent Western Blot Stripping Buffer enables the reuse of PVDF membranes, simplifying the Western blot optimization process and allowing the same blot to be reprobed with different primary antibodies to detect alternative targets. Restore Fluorescent Western Blot Stripping Buffer is for use with low-fluorescence PVDF membrane only (Part No. 22860).

Fluorescence Western blotting is a powerful method for detecting multiple targets at once. Restore Fluorescent Western Blot Stripping Buffer allows re-probing of PVDF membranes, saving time and cost. This is ideal when samples are limited and optimization or analysis with different primary antibodies is required.

Traditional stripping methods may adversely alter or remove the sample proteins from the PVDF membrane during the stripping process or may be effective for removing only low-affinity antibodies. In contrast, Restore Fluorescent Western Blot Stripping Buffer Stripping efficiency exceeds 90% while reprobing efficiency matches or exceeds other supplier's formulations. Also, Restore Fluorescent Western Blot Stripping Buffer is conveniently stored at room temperature and easy to use. Simply dilute buffer 1:5 in water and incubate your membrane for 5 to 20 minutes at room temperature.

WesternBreeze™ Blocker/Diluent (Part A and B) (Invitrogen™)

The WesternBreeze® Blocker/Diluent (part A and B) is an optimized, easy-to-use blocker and primary antibody diluent system that yields low background/high signal western blot detection on nitrocellulose (NC) and polyvinylidene difluoride (PVDF) membranes. Sufficient reagents for 20 mini-blots.

Restore™ PLUS Western Blot Stripping Buffer (Thermo Scientific™)

Thermo Scientific Restore PLUS Western Blot Stripping Buffer is an advanced formula for removing bound primary and secondary antibodies from membranes so they can be reprobed and detected with chemiluminescent substrates.

Features of Restore PLUS Western Blot Stripping Buffer:

Ready and easy to use—no dilution necessary; no offensive odors; store at room temperature
Compatible—use on nitrocellulose and PVDF membranes, whether still wet or already dry; works with practically any blocking buffer, enzyme conjugate and chemiluminescent substrate
Cost effective—save valuable time and samples; strip blots effectively the first time
Robust yet gentle—transferred proteins remain viable; strip the same blot up to five times
Flexible—strip and reprobe to optimize antibody concentrations or to detect a new antigen with different antibodies

Restore PLUS Buffer is an alternative formulation of the original Thermo Scientific Restore Western Blot Stripping Buffer. Restore PLUS Stripping Buffer was designed for use with antibodies that are difficult to remove from western blots and require longer incubation times or incubation temperatures greater than 22°C with gentler formulations. High-affinity antibodies can be quickly and effectively stripped from western blots at room temperature without removing transferred proteins, thereby allowing multiple reprobes of the target.

Protocol Summary:
• Wash blot to remove chemiluminescent substrate
• Incubate blot in Restore Western Blot Stripping Buffer for 5 to 15 minutes at room temperature
(or incubate at 37°C for high affinity antibodies)
• Remove blot and wash in Wash Buffer
• Block membrane
• Test for sufficient removal of antibodies
• Perform next immunoblot experiment

Membrane Blocking Solution (Invitrogen™)

Membrane Blocking Solution can be used to block nitrocellulose and PVDF membranes, as well as to dilute antibodies or enzyme conjugates. This solution is a Tris buffer containing bovine serum albumin, goat IgG, Tween 20, and a mixture of other components that can reduce non-specific binding. The Proclin used in this reagent does not inhibit peroxidase or alkaline phosphatase activity. Note that this blocking solution contains normal goat IgG and should not be used in assays where anti-goat immunoglobulins are employed.

WesternBreeze™ Wash Solution (16X) (Invitrogen™)

WesternBreeze® Wash Solution (16X) is an optimized solution of concentrated buffered saline containing detergent to minimize background and non-specific binding on nitrocellulose (NC) and polyvinylidene difluoride (PVDF) membranes. Sufficient reagents for 20 mini-blots.

Pierce™ Western Blot Signal Enhancer (Thermo Scientific™)

Thermo Scientific Pierce Western Blot Signal Enhancer is a two-reagent system for conditioning protein blots after transfer to greatly enhance the effectiveness of primary antibodies and intensify the final detection signal in Western blot experiments.

Features of Western Blot Signal Enhancer:

Increases protein detection—most protein targets show a three- to 10-fold increase in signal intensity, enabling much less protein to be detected with the same substrate and method
Improves antibody binding—the membrane-treatment reagent exposes and conditions target proteins so that specific antibodies can bind more effectively
Works for nearly any protein—signal enhancement has been demonstrated with targets such as IL-6, p53, NFκB, BRCA1 and EGF
Effective with any substrate—enhances both chemiluminescent (ECL) and colorimetric detection for Western blots
Compatible with any membrane—enhances signal on nitrocellulose and PVDF membrane, regardless of pore size (enhancement is less pronounced with PVDF)
Fast, 15-minute protocol—optimized for a combination of simplicity, speed and signal enhancement for most proteins
Ready-to-use—no formulating or diluting necessary, and the reagents are stable for storage at room temperature

The Pierce Western Blot Signal Enhancer membrane treatment procedure is very simple, takes only 15 minutes and can be added to nearly any existing Western blotting protocol. The result is an increase in the intensity of target protein bands on the Western blot or detection of target proteins at levels that were previously not possible. The product is effective for signal intensification with both chemiluminescent and chromogenic substrates, especially with nitrocellulose membranes.