Solid phase extraction of proteins from buffer solutions employing capillary-channeled polymer (C-CP) fibers as the stationary phase

Abstract

Polypropylene (PP) capillary-channeled polymer (C-CP) fibers are applied for solid phase extraction (SPE) of proteins from aqueous buffer solutions using a micropipette tip-based format. A process was developed in which centrifugation is used as the moving force for solution passage in the loading/washing steps instead of the previously employed manual aspiration. The complete procedure requires ∼15 minutes, with the number of samples run in parallel limited only by the capacity of the centrifuge. The method performance was evaluated based on adsorption and elution characteristics of several proteins (cytochrome c, lysozyme, myoglobin, and glucose oxidase) from 150 mM phosphate buffered saline (PBS) solutions. Protein concentration ranges of ∼2 to 100 μg mL⁻¹ were employed and the recovery characteristics determined through UV-Vis absorbance spectrophotometry for protein quantification. The protein loading capacities across the range of proteins was ∼1.5 μg for the 5 mg fiber tips. Average recoveries from PBS were determined for each protein sample; cytochrome c ∼86%, lysozyme ∼80%, myoglobin ∼86%, and glucose oxidase ∼89%. Recoveries from more complex matrices, synthetic urine and synthetic saliva, were determined to be ∼90%. A 10× dilution study for a fixed 1 μg protein application yielded 94 ± 3.2% recoveries. The C-CP tips provided significantly higher recoveries for myoglobin in a 150 mM PBS matrix in comparison to a commercially available protein SPE product, with the added advantages of low cost, rapid processing, and reusability.