Removal of sodium dodecyl sulfate from protein samples prior to matrix‐assisted laser desorption/ionization mass spectrometry

Abstract

Sodium dodecyl sulfate (SDS) is widely used for protein solubilization and for separation of proteins by SDS polyacrylamide gel electrophoresis (SDS-PAGE). However, SDS interferes with other techniques used for characterization of proteins, such as mass spectrometry (MS) and amino acid sequencing. In this paper, we have compared three procedures to remove SDS from proteins, including chloroform/methanol/water extraction (C/M/W), cold acetone extraction and desalting columns, in order to find a rapid and reproducible procedure that provides sufficient reduction of SDS and high recovery rates for proteins prior to matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS). A 1000-fold reduction of SDS concentration and a protein recovery at approximately 50% were obtained with the C/M/W procedure. The cold acetone procedure gave a 100-fold reduction of SDS and a protein recovery of approximately 80%. By using desalting columns, the removal of SDS was 100-fold, with a protein recovery of nearly 50%. Both the C/M/W and the cold acetone methods provided sufficient reduction of SDS, high recovery rates of protein and allowed the acquisition of MALDI spectra. The use of n-octyl-beta-D-glucopyranoside in the protein sample preparation enhanced the MALDI signal for protein samples containing more than 2 10(-4)% SDS, after the C/M/W extraction. Following the cold acetone procedure, the use of n-octylglucoside was found to be necessary in order to obtain spectra, but they were of lower quality than those obtained with the C/M/W method, probably due to higher residual amounts of SDS.

Application of MS-Based Proteomics to Study Serum Protein Adsorption/Absorption and Complement C3 Activation on Poly(ethylene glycol) Hydrogels

“…Serum proteins adsorbed/absorbed to PEG hydrogels and TCPS were eluted with sodium dodecyl sulfate (SDS, Amresco ® ) or 30% (w/v) acetic acid and directly dried with a SpeedVac concentrator (Savant SVC 100H, Phoenix Equipment). SDS in protein solution was removed by chloroform/methanol/water extraction (1:4:3, v/v) as described previously [17]. Briefly, 200 μl protein sample was mixed with 800 μl methanol, followed by the addition of 200 μl chloroform and 600 μl water.…”

Electroelution without gel sectioning of proteins from sodium dodecyl sulfate-polyacrylamide gel electrophoresis: Fluorescent detection, recovery, isoelectric focusing and matrix assisted laser desorption/ionization-time of flight of the electroeluate

“…In the case of electroelution, SDS can be effectively removed during the elution procedure if a SDS-free elution buffer is used (e.g., [9,[13][14][15][16]). Once proteins are eluted from a gel, a number of procedures can be employed to remove SDS (see, e.g., [18] and references therein). Once proteins are eluted from a gel, a number of procedures can be employed to remove SDS (see, e.g., [18] and references therein).…”

“…Once proteins are eluted from a gel, a number of procedures can be employed to remove SDS (see, e.g., [18] and references therein). SDS can be removed by treating the protein solution with organic solvents (e.g., [18]) or by using chromatographic procedures [18,21]. SDS can be removed by treating the protein solution with organic solvents (e.g., [18]) or by using chromatographic procedures [18,21].…”

“…5, panel A vs. B). This decline in sensitivity cannot be entirely attributed to the residual SDS in the protein sample, which is known to reduce the signal in MS (e.g., [9,18]). The sensitivity of detection has also declined sharply (Section 3.5).…”

Development and application of wide-range gradient gel electrophoresis to proteome analysis

“…[11][12][13][14][15][16][17][18] Although these methods can eliminate SDS and other salts from the protein samples to various degrees, they have their inherent limitations, including signicant protein loss, and therefore are not suitable for the treatment of all kinds of protein mixtures particularly those in micro-amounts. [11][12][13][14][15][16][17][18] Although these methods can eliminate SDS and other salts from the protein samples to various degrees, they have their inherent limitations, including signicant protein loss, and therefore are not suitable for the treatment of all kinds of protein mixtures particularly those in micro-amounts.…”