In situ biological samples deproteinization in microfluidic paper-based analytical devices

Deproteinization by protein precipitation is one of the most frequently used biological samples preparation method. It usually involves the addition of protein precipitating agent to liquid sample, followed by centrifugation. The resulting supernatant is then subjected to further analytical steps. In case of on-site analyses, all analytical steps, including sample preparation, ideally should be executed in a single sensing device. Here, a detailed study of blood serum samples deproteinization in situ in microfluidic paper-based analytical devices (mu PADs) is presented. The idea of this process was to precipitate protein with a precipitating agent immobilized on paper, and owing to filtrating properties of paper and capillary fluid flow, obtaining a protein-free sample in a neighboring zone. Firstly, an optimal protein precipitating agent to be used in mu PADs was selected basing on its effectiveness, long-term stability and versatility in terms of initial protein concentration. Paper-based deproteinization effectiveness was determined for both for artificial protein solutions as well as serum samples. Protein precipitation on paper was visualized with scanning electron microscopy in low vacuum mode. Finally, paper-based sample deproteinization was combined with colorimetric copper(II) ions determination process in a single proof-of-concept mu PAD. Cu(II) recovery in spiked control serum samples was assayed to validate the developed paper-based analytical devices.