All-in-one paper-based sampling chip for targeted protein analysis

Highlights: • All-in-one paper-based sampling concept in chip format for mass spectrometric bottom-up protein analysis. • A simple sampling concept for sensitive analysis of lower abundant proteins related to disease state protein applications. • Paper-based sampling chip integrated with mAb E27 and all steps necessary for analysis of proteins in biological matrices. • A high degree of performance sampled with human serum and freshly collected whole blood samples. • Demonstrated with the biomarker human chorionic gonadotropin. -- Abstract: A novel all-in-one paper-based sampling concept for mass spectrometric bottom-up protein analysis is here demonstrated in a chip format integrating instant immunocapture, protein reduction, - alkylation and tryptic digestion all in-device. Conventional laboratory grade filter paper was coated with the polymer 2-hydroxyethyl methacrylate-co-2-vinyl-4,4-dimethyl azlactone (pHEMA-VDM) with a subsequent covalent immobilization of the monoclonal antibody E27 targeting the biomarker human chorionic gonadotropin (hCG). In-device protein reduction and alkylation was optimized with regards to reagent concentration and reaction pH. The sampling concept showed a high degree of performance between 10 and 1000 ng/mL (R² > 0.99) by a five-point calibration curve sampled with hCG spiked to human serum samples and freshly collected whole blood samples, respectively. LOD (experimentally obtained at 100 pg/mL (2.64 pM/0.9 IU/L)) was demonstrated to be up to ten times lower with more than six times faster sample preparation than what has previously been reported for on-paper analysis of hCG in human serum samples.