DSP precipitates from solution containing less than 10-20% DMSO. I need alternatives since my cells are sensitive to such concentrations of DMSO.
Crosslinkers often have a hydrophobic spacer arm and have to be dissolved in DMSO. Could you switch with a water soluble crosslinker ? you might try sulfo-NHS.
Even 1% DMSO is normally toxic to many type of cells. In our lab, we never go beyond a final concentration of 1/1000 DMSO in cell culture with our drug dilutions, Even 1/1000 DMSO is slightly toxic to cells. I ve worked with Prostate, Testicular, Renal, Breast, Pancreatic and Bladder cancer cell lines and with all of the cell lines I experienced acute toxicity when I had 1/100 DMSO (V/V) as a final conc. in the culture medium.
you can dissolve DSP in an organic solvent, I think it doesn’t necessarily have to be a DMSO.You might be able to use other (less toxic) organic solvents to do this. NaOH, Ethanol or combination of both.
There is also an alternative that you can use which is called DTSSP and is a water-soluble analog of DSP.
DSP is non-sulfonated and, therefore, is non-water-soluble whereas DTSSP is sulfonated and water-soluble. Because DSP does not possess a charged group, it is lipophilic and membrane-permeable and is useful for intracellular and intramembrane conjugation. DTSSP is water-soluble and can be dissolved in water, buffer or added directly to an aqueous media therefore, it is useful for crosslinking cell surface proteins. If you are crosslinking cell surface proteins you may want to consider using DTSSP but if your target is intracellular then, you need to use DSP and try a less toxic organic solvent to dissolve it. Hope you can manage to dissolve it in alternative solvents as 10% DMSO is extremely toxic for any type of cells. hope this helps.
Thank you Knut, Remy and Ahmet for your helpful answers. Ahmet, my target protein is intracellular, so DTSSP won't do. I hadn't considered NaOH or EtOH - Ill try them for resuspension and see how I go. Remy, Ill look into sulfo-NHS - is it cleavable?. I failed to mention that I require a cleavable crosslinker. Thanks again to you all for your help! Very much appreciated.
Hi Corey,
You could use DTBP or DST. Both are cleavable as well as water soluble.
Sulfo-NHS esters provide crosslinkers with water solubility, but they also inhibit the crosslinker from passing the cell membrane. Have you tried DMF instead of DMSO? The following are cell membrane permeable, but would first require dissolving the crosslinker in a water miscible organic solvent:
DSP (http://www.proteochem.com/dspcrosslinker100mg-p-96.html)
DST (http://www.proteochem.com/dstcrosslinker100mg-p-205.html)
EGS (http://www.proteochem.com/egscrosslinker100mg-p-153.html)
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