Protein Crosslinking

Crosslinkers, also known as bifunctional crosslinkers, are reagents that contain two or more reactive groups which covalently attach via a spacer to functional groups on proteins or other biomolecules. Three types of crosslinkers are available: homobifunctional crosslinkers, heterobifunctional crosslinkers, and photoreactive crosslinkers. Homobifunctional crosslinking reagents have identical reactive groups, primarily amine-to-amine or sulfhydryl-to-sulfhydryl. They are typically used to form intramolecular crosslinks or to prepare polymers from monomers. Heterobifunctional crosslinking reagents have different reactive groups such as amine-to-sulfhydryl, carboxyl-to-amine, or sulfhydryl-to-carboxyl. They are useful for preparing conjugates between two different biomolecules. Heterobifunctional crosslinking reagents also include photoreactive crosslinking reagents that react with nucleophiles or form C-H insertion sites after exposure to UV light. Learn more about crosslinking reagents.

Find the optimal crosslinking reagent for your application using our Crosslinker Selection Tool or select from the most common crosslinkers below.

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Protein crosslinking no-weigh packaging formats offer quick and convenient ready-to-use solutions. This format eliminates the need to weigh out small amounts of dry compounds and eliminates the waste of unused material. Simply reconstitute and the reagent is ready to use at the desired concentration.

Protein crosslinking premium grade reagents are high-quality reagents formulated for applications where product integrity and risk minimalization are essential. These premium grade reagents are ideal for long-term research projects that require consistent performance. Crosslinker premium grade products provide quality assurance review, lot sample retention, change control notification, along with an extra level of analytical testing and chemical characterization.

In vivo crosslinking can be used to characterize protein interactions and ligand-receptor interactions irrespective of treatment conditions. Various sizes of in vivo crosslinkers are available to target surface and intracellular proteins for analysis by different methods such as immunoprecipitation (IP), Co-IP, chromatin immunoprecipitation (ChIP), electrophoresis mobility shift assay (EMSA), western blot, immunofluorescence (IF), and immunohistochemistry (IHC).

NHS-ester crosslinkers are widely used for in vivo crosslinking. NHS-ester, amine-specific reactions are fast and highly efficient since lysine residues are abundant and easily accessible on the surface of most proteins. Homobifunctional crosslinkers use a “shotgun” type approach for identifying protein complexes since they crosslink all interacting molecules whose lysine residues are within the crosslinkers spacer length. Subsequent detection simply requires a specific antibody or probe that targets one of the molecules in the complex.

Find homobifunctional, amine-reactive, NHS-ester crosslinkers for in vivo crosslinking

NHS-ester/diazirine crosslinkers combine NHS-ester chemistry with diazirine-based photochemistry to create crosslinking. While typical heterobifunctional crosslinkers require specific amino acid groups at correct distances, SDA crosslinkers utilize a “two-step” reaction by crosslinking one protein using the NHS-ester then UV light to activate crosslinking via the diazirine moiety to any amino acid on a second protein.

Find heterobifunctional, photoreactive, NHS-ester/diazirine crosslinkers for in vivo crosslinking

Photo-Leucine and Photo-Methionine, used in metabolic labeling, are incorporated into proteins by protein synthesis (3). These non-toxic photo-amine acids are non-invasive and do not alter the cell’s metabolism. Photo-Leucine and Photo-Methionine are highly specific and ideal for studying hydrophobic interactions. Further, they can be used for either extracellular or intracellular crosslinking.

Find photoreactive amino acid crosslinkers for in vivo crosslinking

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