Capillary-channeled polymer (C-CP) fibers for the rapid extraction of proteins from urine matrices prior to detection with MALDI-MS

Abstract

Purpose: While MS is a powerful tool for biomarker determinations, the high salt content and the small molecules present in urine poses incredible challenges. Separation/extraction methods must be employed for the isolation of target species at relevant concentrations. Micropipette tips packed with capillary-channeled polymer (C-CP) fibers are employed for the SPE of proteins from a synthetic and a certified urine matrix.

Experimental design: Extractions are performed utilizing a very simple centrifugation method to spin-down species through the C-CP fiber tips. Proteins adsorb to the hydrophobic polypropylene fibers and are eluted in a solvent suitable for MALDI-MS analysis. Figures of merit are determined for representative compounds β2-microglobulin, retinol binding protein, and transferrin.

Results: The optimum protein processing included a 100 μL aqueous rinse and an elution solvent composition was 10 μL of 55:45 ACN:water (with triflouroacetic acid). MALDI-MS responses for the target proteins are improved from nondetectable levels to eventually yield LOD ranging from 5 to 180 nM in 1 μL aliquots.

Conclusion and clinical relevance: C-CP fiber tips offer a plethora of advantages including low materials costs, high throughput, microvolume processing, and the determination of sub-nanogram quantities of analyte; allowing determination of biomarkers that are otherwise undetectable in urine matrices.

Keywords: Capillary-channeled polymer fibers; MALDI-MS; Proteins; Solid-phase extraction; Urine.