Click Chemistry Reagents from LI-COR® for Biomolecule Labeling

Biomolecule labeling continues to be a cornerstone feature of many in vitro and in vivo biological experiments. Click Chemistry has recently emerged as a convenient, versatile, and reliable method for labeling a wide variety of molecules for applications ranging from biomarker isolation to assay development.
Click Chemistry Workflow
LI-COR now offers a portfolio of Click Chemistry reagents for copper-catalyzed and copper-free methods. These products offer researchers flexibility to choose the correct reagent for a diverse array of applications. LI-COR Click Chemistry reagents include IRDye® 800CW, IRDye 680RD, and IRDye 650 infrared fluorescent dyes labeled with DBCO, azide, or alkyne groups.

Click Chemistry utilizes pairs of reagents that exclusively react with each other and are effectively inert to naturally-occurring functional groups such as amines. Unlike affinity interactions such as streptavidin-biotin, Click Chemistry forges covalent bonds between the reacting partners to deliver stable bioconjugates.

Click Chemistry reactions can be categorized into two separate groups, copper-catalyzed or copper-free. Copper-catalyzed Click Chemistry is used for initiating reactions between azides and alkynes. These reactions are also known as Copper-Catalyzed Azide-Alkyne Cycloaddition (CuAAC). Although they initiate and accelerate Click Reactions, copper catalysts are cytotoxic and inappropriate for use in living systems.

Watch this informative webinar on IRDye Infrared Dye Reagents for Click Chemistry.

NEW! IRDye® Goat Anti-Mouse IgM Secondary Antibodies from LI-COR®!

IRDye Dye-labeled Goat anti-Mouse AntibodiesOur IRDye secondary antibody line is growing! We have recently added IRDye Goat anti-Mouse IgM (μ chain specific) secondaries labeled with:

  • IRDye 800CW (PN 926-32280)
  • IRDye 680RD (PN 926-68180) or
  • IRDye 680LT (PN 926-68080).

Just like all of the LI-COR IRDye secondary antibodies, these are highly cross-adsorbed secondary antibody conjugates suitable for a variety of applications (see the table below).

IRDye 800CW secondary antibodies are the antibodies of choice for a wide variety of applications in the 800 nm channel (see the list below). IRDye 800CW secondary antibodies can be used for 2-color detection when multiplexed with IRDye 680RD or IRDye 680LT secondary antibodies.

IRDye 680RD secondary antibodies are the antibodies of choice for In-Cell Western Assay and Western blot applications in the 700 nm channel. These antibodies can be used for 2-color detection when multiplexed with IRDye 800CW secondary antibodies. These antibodies are our most universal use 700 nm channel antibodies. Start using IRDye 680RD first over other 700 nm dyes. Dilution working range 1:10,000 – 1:40,000.

IRDye 680LT secondary antibodies have been proven the brightest signal for Western blot detection in the 700 nm channel and are comparable to Alexa Fluor 680 secondary antibodies. Choose IRDye 680LT secondary antibodies to get high signal and for specific uses of detection in the 700nm channel. These antibodies are not recommended when getting up and running on system. Once established near-infrared protocols are optimized with IRDye 680RD, IRDye 680LT can be used to optimize signals in the 700 channel. Dilution range 1:20,000 – 1:40,000. Note: optimization may be required with IRDye 680LT.

Application IRDye 800CW
IRDye 680RD
IRDye 680LT
Western Blot
In-Cell Western™ Assay Not Recommended
On-Cell Western Assay Not Recommended
Protein Array
2D Gel Detection
Tissue Section Imaging
Small Animal Imaging Not Recommended
Virus Titration Assay Not Known Not Known
FRET-based Assay Not Known Not Known

Note: Now, as of December 15, 2014, you can also get 0.1 mg sizes of all of our IRDye dye-labeled secondary antibodies. Check out our complete listing here and our new filtering tool!

Fast, Cost-Effective Cell-Based Assays for Quantitative Cell Signaling Analysis

In-Cell Western Assay Kits for Quantitative Cell Signaling Analysis
The In-Cell Western™ Assay is an immunocytochemical assay that uses near-infrared fluorescence to detect and quantify proteins in fixed cells. Detecting proteins in their cellular context increases quantification precision. Proteins in fixed, cultured cells are detected directly in microplates, which yields higher throughput compared to Western blotting and eliminates typical Western blotting steps such as cell lysate preparation, electrophoresis, and membrane transfer. Using the In-Cell Western Assay kits, the cost per well for secondary screening is reduced to a fraction of the cost of typical screening methods. Watch an introductory webinar to In-Cell Western Assays.

The CellTag™ 700 Stain ICW Kits provide antibodies, blocking buffer, and CellTag 700 Stain to normalize well-to-well variations in cell number for forty 96-well plates or ten 384-well plates. Using protein stains reduces the cost per assay compared to performing the assay using two secondary antibodies. Any potential interference caused by using two antibodies is also eliminated.
Phospho-ERK Analysis with Sapphire700/DRAQ5 for Normalization

Additional Resources:

Other In-Cell Western Assay Protocols are available on our Technical Resources Library. Visit our In-Cell Western Application Pages for more examples of how researchers have used these immunocytochemical assays.

Updated February 18, 2015.

Now Use the Same Primary Antibody for Immunoprecipitation AND Western Blotting!

Quick Western Kit - IRDye 680RD, PN 926-68100

LI-COR® has just rolled out a new way that the recently-released Quick Western Kit – IRDye® 680RD (see Would you like to save at least 90 minutes the next time you do a Western blot?) can be used in your research.

Not only can the Quick Western Kit reduce Western blotting time by 90 min, the kit ALSO serves as a detection solution for post-immunoprecipitation samples by Western blot because it does not bind to denatured mouse monoclonal or rabbit monoclonal antibodies. The key benefit is the ability to use the same antibody for immunoprecipitation and post-immunoprecipitation detection by Western blot. Seriously, how cool is that!!??!!

Using the same primary antibody for IP and Western blotting with the Quick Western Kit

Figure 1. A431 cell lysates were immunoprecipitated overnight with a monoclonal antibody against p53. The resulting immunoprecipitates were separated by SDS-PAGE. Lane 1: Negative IP control; Lane 2: Test sample ; Lane 3: A431 cell lysate positive control. Western blotting was performed using the same p53 monoclonal antibody and incubated with IRDye 680RD Immunoprecipitation Detection Reagent.

Protocol: Detection of post-immunoprecipitation proteins by Western blot using the Quick Western Kit – IRDye 680RD

For more information, visit our website. Here’s the kit pack insert. To order this product (online ordering available in select countries), go to our ecommerce site.

Money-Saving Quick Western Kit Reduces Western Blotting Time by at Least 90 min

Introducing the NEW! Quick Western Kit – IRDye® 680RD

Quick Western Kit - IRDye 680RD

The Quick Western Kit – IRDye® 680RD provides a universal detection reagent that can be combined with the primary antibody incubation step, eliminating the need for a secondary antibody incubation step.

Saves Time (View Workflow)

  • Does not require a separate primary antibody labeling step, saving time and antibody
  • Faster method of detection compared to the traditional 2-step method, which can take up to 4 hours
  • Reduces the total Western blot procedure by at least 90 min

The kit can be used to detect primary antibodies from a variety of hosts and has been shown to recognize primary antibodies to recombinant tagged proteins (i.e. 6X His, Myc, DDK, etc.)

IRDye 680RD Detection Reagent is known to have high specificity for IgG from:

  • Human
  • Mouse
  • Rabbit
  • Guinea Pig
  • Goat
  • Sheep
  • Pig
  • Cow
  • Cat
  • Dog
  • Donkey

The IRDye 680RD Detection Reagent does not work with:

  • Chicken

The Detection Reagent is known to have lower specificity for rat, horse, and hamster. The Detection Reagent has been specifically tested and qualified for Western blot applications. If additional specificity and/or affinity are required, please use IRDye conjugated secondary antibodies for detection.
Quick Western Kit data

Two-fold dilutions of crude lysate containing an overexpressed 6X-His tagged DNA polymerase was loaded in Lanes 3-7. A Histag molecular weight marker (Invitrogen) was loaded in Lanes 1 & 9. The nitrocellulose membrane was blocked with Odyssey Blocking Buffer (PBS) and probed with anti-His Tag Rabbit Polyclonal Antibody (GenScript; 1:500) and IRDye 680RD Detection Reagent (1:1000) for 1 hour. The image was collected on the Odyssey CLx.

Ordering Information
Pack Insert

New Tools for Cancer Surgeons: Targeted Fluorescent Imaging Probes

LI-COR interviewed Dr. Go van Dam, a surgeon specializing in oncology at the Groningen University Medical Center in the Netherlands.

A key focus of van Dam’s research is to explore new tools such as targeted fluorescent imaging probes that will help address the challenges facing oncology surgeons. He discusses his research using near-infrared fluorescent imaging during surgery to improve cancer patient outcomes. Watch an interview with Dr. van Dam.

Vasilis Ntziachristos, PhD,  Technische Universität München, Germany and Gooitzen M. van Dam, MD, PhD, University Medical Center Groningen, Netherlands presented “Shining New Light on Clinical Fluorescence Imaging” at World Molecular Congress in San Diego, CA in September 2011.