Western Blot Stripping Buffer (Neutral)

Introduction

Western Blot Stripping Buffer is used for the reuse of the transferred protein membrane in Western blot. After primary antibody binding to the membrane and secondary antibody binding to the primary antibody, then using ECL subsequent to exposure the images, if the samples on the membrane are also needed to verify some control proteins or other target proteins, this product can be used to elute the combined antibodies before. After the antibody signals are cleared, the membrane can be re-incubated with control antibodies or other antibodies for the second verification. It can save the time of preparing SDS-PAGE gel and transfer, and the samples of experiment are also saved.

It only takes about 30 min to reuse the protein membrane which can eliminate the error caused by redoing the experiment and makes the experiment results more comparable.

Components

Experimental Procedure

1. After doing the first Western blot experiment, wash the membrane with TBST buffer for 5 min and repeat 3 times.

2. Take the membrane to a tank with Western Blot Stripping Buffer in it, make sure that the membrane is completely immersed in the Western Blot Stripping Buffer, gently shake the tank to elute the antibodies at room temperature for 30 min.

3. Discard the Western Blot Stripping Buffer and wash the membrane with TBST buffer for 5 min and repeat 3 times.

4. (Optional)

1)  Check whether the antibodies have been cleared, block the PVDF membrane and incubate with secondary antibody, then exposed with ECL to observe the signal.

2)  If the antibodies have been cleared, clean the PVDF membrane with TBST buffer for 5 min and repeat 3 times. Block the membrane and incubate with a new primary (or Control) antibody.

3)  If there is any signal on the membrane, follow the step "2" and extend the elution time until the signals are cleaned out.

5.  Block the membrane and incubate with a new primary (or Control) antibody according to the Western blot experimental procedure.

Storage

Store at 2~8°C for 12 months.

Cautions

1. When the PVDF membrane is immersed with this product, the membrane will become transparent, which is a normal phenomenon.

2. In order to obtain the best performance, PVDF membrane is recommended for this product.

3. This product is recommended to ECL or similar chemiluminescent reagents. If non-chemiluminescent reagents, such as DAB is used to expose the image, please don’t use this product.

4. Repeated use of the same membrane with western blot experiments may lead to the weakening of protein signal. The membrane can be reused 3~5 times with several antibodies tests.

5. Repeated washing the PVDF membrane may lead to weak signal, please detect the lower expression protein first, and higher proteins such as control after cleaning with Western Blot Stripping Buffer.

6. For your safety and health, please wear experimental clothes and disposable gloves.