Blocking Buffer Choice Is Important for Western Blot Success

Blocking Buffers


LI-COR offers several blocking buffers for use with the Odyssey® Imaging Systems. Intercept® Blocking Buffers are optimized for use with IRDye® products and other near-infrared fluorophores. They provide excellent performance with low variability and low background for quantitative Western blots and other immunoassays.

Because no single blocking buffer is ideal for every experiment, several varieties of Intercept Blocking Buffer are available.

Intercept Blocking Buffers can be used to dilute HRP-conjugated secondary antibodies and can be used in the blocking step prior to incubating the blot with HRP-conjugated secondary antibodies. Because Intercept Blocking Buffers contain no sodium azide, they can provide the same excellent performance for chemiluminescent Western blot detection, as well.

Choose the Right Blocking Buffer for Your Application

Blocking buffer choice is important for immunoassay success. Blockers enhance sensitivity by reducing background interference, increasing signal-to-noise ratio, and promoting specific binding of the primary antibody while minimizing non-specific interactions. Choosing the right blocker is important because:

  • Insufficient blocking yields high background and reduced signal-to-noise ratios.
  • Excessive blocking may cause loss of blotted proteins1 or mask antibody:antigen interactions.
  • No single blocking buffer selection is suitable for ALL antigen-antibody pairs.
  • Blocker choice may affect antibody specificity and non-specific binding.2

Several options of Intercept Blocking Buffer are available to provide one that best suits your experiment.

Intercept Protein-Based Blocking Buffers

Intercept non-mammalian protein-based blocking buffers provide the same highly efficient blocking with high signal-to-noise ratio in immunoassays as Odyssey Blocking Buffers, but in a new, cost-effective formula.

With a convenient 1x formulation, you won’t need to dilute Intercept Blocking Buffers. Intercept Blocking Buffer is ready to use, so you can block and move on with your research.

Figure 1: Western blot protocol demonstrating the comparison of Intercept Blocking Buffer to Odyssey Blocking Buffer. Blots with serial dilution of mouse heart tissue extract were probed for Stat3 Mouse mAb and p38 MAPK Rabbit Ab, detected with IRDye 800CW Goat anti-Mouse IgG and IRDye 680LT Goat anti-Rabbit IgG, and scanned together on an Odyssey CLx Imaging System.

Intercept Protein-Free Blocking Buffers

Protein-free blocking buffers are essential for applications where animal-based products are prohibited or protein-based blocking buffers are contributing to cross-reactivity. Designed for quantitative and chemiluminescent Western blots imaged on an Odyssey Imager, Intercept Protein-Free Blocking Buffers provide an effective and budget-friendly alternative to protein-based blocking buffers.

Note: Intercept Protein-Free Blocking Buffers now use an eco-friendly ingredient. While this ingredient changes the appearance of the blocking buffer, the blocking buffer still performs the same.

LI-COR Blocking Buffers can be used in many applications, including:

man browsing man browsing man browsing

Blocking Buffers for Western Blotting

Sign up to review the science, best practices, and mechanics of the blocking buffer step for either chemiluminescent or near-infrared fluorescent Western blots at Lambda U® online learning portal.

These online, on-demand resources can help ensure that you are getting the very best Western blot data.

Sign Up


  1. DenHollander, N., and Befus, D. Journal of Immunological Methods, 122(1), pp 129-135. DOI: 10.1016/0022-1759(89)90343-8.
  2. Ambroz, K.L., Zhang, Y., Schutz-Geschwender, A., and Olive, D.M. (2008). Protemoics, 8(12), pp 2379-2383. DOI: 10.1002/pmic.200700676.