Can you provide the shelf-life for DSG (disuccinimidyl glutarate)?
DSG (disuccinimidyl glutarate) is covered under our general 1-year warranty and is guaranteed to be fully functional for 12 months from the date of shipment, if stored as recommended. Please see section 8.1 of our Terms & Conditions of Sale (https://www.thermofisher.com/content/dam/LifeTech/Documents/PDFs/Terms-and-Conditions-of-Sale.pdf) for more details.
Find additional tips, troubleshooting help, and resources within our Protein Purification and Isolation Support Center.
How should I dissolve and store DSG, No-Weigh Format?
DSG is an NHS-ester that hydrolyzes easily and is moisture-sensitive. For optimal results, we recommend storing DSG powder desiccated at 4 degrees C. To avoid moisture condensation onto the product, the vial must be equilibrated to room temperature before opening. We recommend resuspending DSG in DMSO, Anhydrous (Cat. No. D12345) right before use and not storing the solution.
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How does DSP work as a crosslinker?
DSP is a homobifunctional, water soluble, cleavable, amine-reactive cross-linker. The amine-reactive ends are NHS-esters. This group reacts with primary amines, in proteins that translates to lysine groups and N-terminus amines. It reacts best in the pH range of 7.5 to 8.5, this ensures the amines are deprotonated and it limits the hydrolysis of the ester at higher pHs. NHS-esters are best used in a heavy molar excess over the number of amines present. This helps to overcome the competing hydrolysis of the ester to an inactive form.
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How is the DSP cross-linking reaction quenched?
1M Tris (or glycine), pH 7.5 can be added to the reaction mixture to a final concentration of approximately 20-50 mM.
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How is DSP cleaved?
The disulfIde bond in DSP's 12Å spacer arm can be cleaved by at least three different methods. The first is by incubating the crosslinker in 10-50 mM DTT for 30 minutes at 37 degrees C. Secondly, the disulfIde can be reduced by incubation in 5% 2-Mercaptoethanol (2-ME) in SDS-PAGE sample buffer at 100 degrees C for 5 minutes. Finally, the “-S-S-” bond can be cleaved by incubation in 100 mM sodium borohydride at room temperature for 30-60 minutes. Other reducing agents such as TCEP-HCl or 2-Mercaptoethylamine (2-MEA) should also work, however these applications have not been referenced.
Find additional tips, troubleshooting help, and resources within our Protein Purification and Isolation Support Center.
