FAQs about PPS Silent® Surfactant

Basics | Properties | Reaction Conditions | Chromatography | Gels and CE

Basics

1) What is PPS Silent Surfactant? PPS Silent Surfactant is a mass spectrometry compatible reagent for extraction and solubilization of hydrophobic proteins and improvement of in-solution enzymatic protein digestions.

2) What does PPS stand for? PPS Silent Surfactant is a pyridinium propyl sulfonate.

3) What is the structural formula of PPS?
3-[3-(1,1-bisalkyloxyethyl)pyridin-1-yl]propane-1-sulfonate.
The structure of PPS is shown below:

4) Does PPS Silent Surfactant affect mass spectrometry? PPS is specifically designed to have minimal negative impact on mass spectrometry. Reducing the pH of the digest buffer cleaves this reagent, allowing MALDI-TOF MS, HPLC/MS, or HPLC/UV analysis without detergent interference.

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Properties

1) What is the CMC of PPS Silent Surfactant? The CMC is 0.128 mM.

2) Is PPS Silent Surfactant hygroscopic? Yes. For best results, follow the product insert Use and Storage Instructions.

3) Is PPS Silent Surfactant ionic, non-ionic, or zwitterionic? PPS Silent Surfactant is zwitterionic.

4) Does PPS Silent Surfactant dissolve in ordinary buffers (e.g. Tris or Ammonium Bicarbonate) and organic solvents? PPS Silent Surfactant is water soluble and will replace most mass spectrometry incompatible detergents in a variety of biological buffers and solvent systems.

5) Can PPS Silent Surfactant be used in place of SDS? PPS Silent Surfactant is zwitterionic, so it cannot be used as a substitute for SDS in gels. However, it can be used in isoelectric focusing. The addition of SDS to focused proteins does not interfere with conventional 2D PAGE analysis.

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Reaction Conditions

1) What pH is required for acid hydrolysis of PPS Silent Surfactant? At neutral pH, the surfactant is stable for many hours; at pH <4 cleavage becomes facile. The lower the pH, the faster cleavage will occur.

2) What acids are used to cleave PPS? Our recommended protocol uses HCl. However, if this condition is not suitable for your application, organic acid such as TFA, formic acid, and acetic acid may be used. The time required for cleavage must be adjusted accordingly (see previous).

3) Is PPS Silent Surfactant trypsin-compatible? Yes. Trypsin is active in the presence of PPS.

4) How much protein can you use per reaction? Up to 10 mg/ml of protein is effectively solubilized by 0.1% PPS Silent Surfactant.

5) Is it OK to use protease inhibitors in combination with PPS Silent Surfactant? We have not closely characterized the effect of protease inhibitors on PPS Silent Surfactant performance. In our experience, the addition of protease inhibitors has not caused any problems.

6) Is it OK to use urea in combination with PPS Silent Surfactant? The addition of urea to a detergent solution may inhibit micelle formation. In most instances, the recommended PPS Silent Surfactant concentration of 0.1% will be effective, even in the presence of urea. If more detergent activity is required, the concentration of PPS Silent Surfactant may be increased to as high as 2%.

7) Is it OK to use PPS Silent Surfactant at high temperature? Yes. Heat may accelerate hydrolytic cleavage of PPS Silent Surfactant in acid conditions.

8) Does PPS Silent Surfactant work for the extraction of organellar proteins? Yes. Our customers have successfully used PPS Silent Surfactant to analyze mitochondrial proteins.

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Chromatography

1) Is the use of PPS Silent Surfactant compatible with reverse-phase chromatography? Yes.

2) Does PPS Silent Surfactant stick to reverse phase column material? The hydrophobic portion of cleaved PPS Silent Surfactant is 1-hexanol, which does not associate strongly with any kind of reverse phase material. Unexpected downstream effects may be caused by the presence of uncleaved surfactant. In this case, cleavage conditions should be optimized by reducing the pH of the digest buffer by one unit or by extending the reaction time interval ten-fold.

3) Is PPS Silent Surfactant compatible with MudPIT? Yes.

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Gels and CE

1) Can PPS Silent Surfactant be used in gels? PPS Silent Surfactant is zwitterionic and therefore cannot be used as a direct replacement for SDS in gels. Proteins solubilized with PPS Silent Surfactant can be run in 2D gels and westerns.

2) Is PPS Silent Surfactant compatible with CE? Yes. Capillary electrophoresis separations may be sensitive to the presence of uncleaved PPS Silent Surfactant. If the end application is CE, good results may require optimizing PPS Silent Surfactant cleavage conditions by reducing the pH of the cleavage reaction solution, or by extending the cleavage reaction time interval.

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