Efficient, Low-Cost, and High-Throughput Sodium Dodecyl Sulfate (SDS) Removal from Protein Digests Using Weak-Anion...

Area of Science:

• Biochemistry
• Analytical Chemistry
• Proteomics

Background:

• Sodium dodecyl sulfate (SDS) is essential for protein denaturation and extraction but interferes with liquid chromatography-tandem mass spectrometry (LC-MS/MS).
• High SDS concentrations cause issues like column overpressure, retention time shifts, and signal suppression in LC-MS/MS.
• Effective SDS removal is critical for reliable LC-MS/MS analysis of protein digests.

Purpose of the Study:

• To develop an inexpensive, high-throughput method for removing SDS from protein digests.
• To validate the efficiency and peptide recovery of the SDS removal method.
• To assess the method's utility in quantifying proteins in complex biological matrices like serum.

Main Methods:

• Utilized weak-anion exchange (WAX) resins in 96-well filter plates for SDS removal.
• Processed protein digests with varying SDS concentrations (0.1-1%).
• Analyzed peptide recovery and quantified trastuzumab in spiked mouse serum post-SDS removal.

Main Results:

• Reduced SDS concentrations from 0.1-0.4% to <5 ppm and 0.6-1% to <100 ppm in under 3 minutes.
• Achieved approximately 90% recovery for unmodified tryptic peptides and ~70% for phosphorylated peptides.
• Demonstrated excellent linearity (R² = 0.9996) and accuracy for trastuzumab quantification in mouse serum.

Conclusions:

• The WAX resin method provides an efficient, cost-effective solution for SDS removal prior to LC-MS/MS.
• This high-throughput technique ensures high peptide recovery and accurate protein quantification.
• The method's affordability and ease of implementation make it highly transferable to other laboratories.