Intercept (TBS) Protein-Free Blocking Buffer

Intercept (PBS) Protein-Free Blocking Buffer

Specifications

• Intercept Protein-Free Blocking Buffers are ready-to-use. For optimal performance, do not dilute.

• Shelf Life: See expiration date on bottle.

• Storage: 4 °C

• Suitable for a variety of applications, including: quantitative Western blots, chemiluminescent Western blots, In‑Cell Western™ Assay, In-Gel Western, protein array, and glycoprotein detection.
• Compatible with avidin-biotin detection systems. Unlike casein or milk-based blocking buffers, Intercept Protein-Free Blocking Buffer will not interfere with avidin-biotin detection systems.

Product Description

Blocking buffers enhance the quality of Western blots by reducing background interference, increasing the signal-to-noise ratio, and promoting specific binding of the primary antibody while minimizing non-specific interactions. Intercept Protein-Free Blocking Buffers are ready-to-use formulations in phosphate-buffered saline (PBS) or tris-buffered saline (TBS) to provide optimal blocking conditions for PBS or TBS systems.

Intercept Protein-Free Blocking Buffer is an excellent alternative when protein-based blocking buffers are contributing to cross-reactivity.

Materials Required for Blocking Membrane

• Blotted PVDF or nitrocellulose membrane

• Western blot incubation box

• Orbital shaker for incubation at room temperature

Guidelines

Use the same buffer (PBS or TBS) throughout the entire Western blot process, including in blocking buffer, antibody diluent, and wash solutions.

Intercept T20 Protein-Free Antibody Diluents contain Tween® 20 and are available in TBS and PBS formulations, so you can use the appropriate buffer for your antibody diluent, without having to spend time measuring and adding Tween 20.

Protocol

For a detailed Western blot protocol, see the Near-Infrared (NIR) Western Blot Detection Protocol (licorbio.com/wbanalysis). For troubleshooting ideas, see Good Westerns Gone Bad: Tips to Make Your NIR Western Blot Great (licorbio.com/GWGBIR).

Blocking Membrane

1. Place the PVDF or nitrocellulose membrane into the Western Blot Incubation Box and add undiluted Intercept Blocking Buffer to the membrane. Be sure to use sufficient blocking buffer to cover the membrane (a minimum of 0.4 mL/cm² is suggested).

   Do not use detergents like Tween® 20 or SDS during the blocking step, as this may generate a background signal. Detergents should only be added when diluting the primary and secondary antibodies.

2. Incubate for 1 hour at room temperature on an orbital shaker.

3. Proceed with Western blot detection protocol.

Creating Antibody Diluent

• Intercept T20 Protein-Free Antibody Diluent contains Tween® 20, so you don't have to spend time measuring and adding Tween 20. PBS and TBS formulations of Intercept T20 Protein-Free Antibody Diluent are available.

• If you're using a PVDF membrane, be sure to add SDS to a final concentration of 0.01 - 0.02% in Intercept Protein-Free Blocking Buffer or Intercept Protein-Free Antibody Diluent when creating secondary antibody diluent.

  See the Near-Infrared Western Blot Protocol (licorbio.com/wbanalysis) for more detail.

Related Products

See licorbio.com/intercept for ordering information.

Blocking Buffer

• Intercept (PBS) Blocking Buffer
• Intercept (TBS) Blocking Buffer

Antibody Diluent

• Intercept T20 (PBS) Antibody Diluent
• Intercept T20 (TBS) Antibody Diluent
• Intercept T20 (PBS) Protein-Free Antibody Diluent
• Intercept T20 (TBS) Protein-Free Antibody Diluent