LI-COR Reagents

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Blocking Buffers

Blocking buffer coats the regions on the blot where there is no protein. It can also be used to dilute antigens, antibodies, and other proteins. Blocking buffer enhances sensitivity by:

  • Reducing background and interference.
  • Increasing signal-to-noise ratio.
  • Minimizing non-specific interactions with primary antibodies.

Blocking Buffers Available from LI-COR

LI-COR offers four varieties of InterceptTM Blocking Buffer: Intercept Blocking Buffer in PBS and TBS, and Intercept Protein Free Blocking Buffer in PBS and TBS. The choice between the two options depends on the experiments being performed. The Intercept Blocking Buffer is optimized for NIR fluorescent assays (NIR Western blot, On-cell Western, etc.). The buffer does not contain azides and can be used for chemiluminescent assays as well.  Intercept™ Blocking Buffer is available in a variety of sizes. 

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Intercept Blocking Buffer (PBS, TBS)

Intercept Blocking Buffer (PBS, TBS) is available in 500 mL bottles and it can be bought in bulk in groups of 3, 10, 25, or 50.

Intercept Protein-Free Blocking Buffer (PBS, TBS)

Intercept Protein-Free Blocking Buffer (PBS, TBS) is available in 500 mL bottles and it can be bought in bulk in groups of 3 or 10.

Western blot protocol demonstrating the comparison of Intercept Blocking Buffer to Odyssey Blocking Buffer. 

Casein Blocking Buffer

Casein Blocking Buffer is available in 100 mL and 500 mL bottles.

Advantages of Intercept Blocking Buffers

  • They are developed and optimized for use on the Odyssey Family instruments.
  • They are rigorously tested to guarantee performance.
  • We are able to help support and troubleshoot any problems that arise using our reagents.
  • They are available in both protein-based and protein-free options, which allows for it to be used in more experiments.

Other types of Blocking Buffer

Researchers sometimes use non-fat milk or Bovine Serum Albumin (BSA) because these are low-cost options. However, their results are sub-optimal because of background illumination and they do not lead to reproducible or robust data.

Caution Against Using Powdered Milk as a Blocking Buffer

Many researchers use powdered milk as a blocking buffer, but this often times is not an ideal blocking buffer. There are several situations where milk should not be used:

  • With anti-goat antibodies - These will cross react with the bovine IgG antibodies found in milk.
  • When using a streptavidin-biotin detection system - Milk contains endogenous levels of biotin.
  • When detecting phosphorylated proteins. - Milk contains casein which is a phospho-protein.

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Antibody Diluents

Intercept T20 Antibody Diluents for use with Odyssey® Imaging Systems, are optimized for use with IRDye® Infrared Dye Reagents and other near-infrared fluorophores.

What are Antibody Diluents?

Antibody diluents are used for the dilution of primary antibodies to the working concentration required for your experiments.

Antibody diluents improve binding affinity of your primary antibody to your antigen. This reduces background due to non-specific binding.

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IRDye ® Secondary Antibodies

Secondary antibodies are used in a Western blot to bind specifically to the primary antibody that is bound to the target. The secondary antibody is labeled with a detection molecule, such as HRP for chemiluminescent detection or a fluorescent dye, such as IRDye 800CW, for NIR detection.

Advantages of NIR Detection

Near infrared detection offers a several advantages over chemiluminescent detection:

  1. It is non-enzymatic and does not change over time.
  2. The signal is directly proportional to the amount of protein on the blot (within the linear range).
  3. It allows for multi-target detection.

Importance of Cross-Adsorption

Cross-adsorption is a purification process where antibodies that may react with a species other than the target are removed from the raw secondary antibody. Choosing high quality secondary antibodies is crucial in order to obtain the best Western blot results. IRDye secondary antibodies are highly cross-adsorbed, which will ensure the best possible two color data.

IRDye Secondary Antibodies Available from LI-COR

LI-COR provides several antibody options for infrared and visible detection.

Infrared antibodies: IRDye 800CW, IRDye 680RD, IRDye 680LT
Visible antibodies: IRDye 650, VRDye 549, VRDye490

IRDye secondary antibodies from result in cleaner blots because they are highly cross-absorbed.

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Chemiluminescence Reagents

Chemiluminescent detection is an enzymatic reaction between luminol, peroxide and HRP. This enzymatic reaction produces light and can be captured using film or a digital imager.

Chemiluminescent Reagents Available from LI-COR

WesternSure reagents are designed and optimized for chemiluminescent research when using film, C-DiGit, Odyssey Fc imaging systems or even other CCD imaging systems. There are several reagents available in this family including WesternSure PREMIUM chemiluminescence substrate, WesternSure HRP-labeled secondary antibodies, WesternSure Pen, WesternSure Pre-stained protein ladder, and WesternSure ECL Stripping Buffer.

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WesternSure PREMIUM Chemiluminescent Substrate 

  • Optimized for use with the C-DiGit and Odyssey Fc Imaging Systems.
  • Dura-like sensitivity for > 60 minutes.
  • Available in 100 mL and 20 mL kits.
  • Can be used with digital imagers and film.

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WesternSure Pen

  • Delivers ink that allows chemiluminescent detection.
  • Can be used to label the lane or mark the bands of the ladder if a non-chemiluminescent ladder was used.
  • Works with film and digital imaging systems.

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WesternSure Pre-stained Chemiluminescent Protein Ladder

  • Has multi-color bands that can be seen on gel and in membrane.
  • Reacts with the substrate in order to provide chemiluminescent signal for digital and film detection.
  • Has 9 bands from 8-260 kDa.
  • Does not rely on strep-tagged HRP or secondary antibody binding.
  • Works with a variety of substrates.

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WesternSure HRP-Conjugated Secondary Antibodies

Available in two configurations: Goat anti-mouse (minimal cross reactivity with human immunoglobulins) and Goat anti-rabbit (minimal cross reactivity with human and mouse immunoglobulins).

Compatible with other chemiluminescent substrates, but optimized for use with WesternSure PREMIUM substrate.

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WesternSure ECL Stripping Buffer

  • Ideal for stripping and re-probing chemiluminescent Western blots.
  • Removes primary and secondary antibodies.
  • Works with both PVDF or Nitrocellulose membranes.
  • Maintains target antigen integrity for efficient re-probing.
  • Does not require hazardous shipping.

WesternSure ECL Stripping Buffer is effective for reprobing with different antibodies

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Western Blot Membranes and Membrane Kits

A low-background membrane is essential for infrared Western blot success. Background can be attributed to membrane autofluorescence or to detection of antibody non-specifically binding to the membrane.  LI-COR have a variety of membranes to suit your western blot needs.

LI-COR offers high-quality, prescreened Immobilon®-FL PVDF, without lot-to-lot variability, packaged in affordable kits with the Intercept Blocking Buffer (PBS or TBS), or with 4X Protein Sample Loading Buffer. LI-COR also offers nitrocellulose membranes in both sheets and rolls.

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Other LI-COR Reagents

LI-COR have an extensive range of reagents and consumable products. Here's a list of other reagents they have to offer.

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