The application of a microfluidic reactor including spontaneously adsorbed trypsin for rapid protein digestion of human tear samples.

ABSTRACT

PURPOSE: The application of a newly developed microfluidic immobilized enzymatic reactor (IMER) designed to accelerate protein digestion in clinical samples is presented. EXPERIMENTAL DESIGN: The IMER contains trypsin adsorbed on the porous surface of a PDMS microfluidic chip. Human tear with its relatively low volume and high protein content is collected and used for testing the digestion efficiency of the IMER. With the use of CZE peptide mapping, the efficiency and reproducibility of the reactor are investigated. RESULTS: No significant difference is observed in the CZE peptide profiles of the same tear sample digested in-solution or via microfluidic IMER. LC-MS measurements show that the microfluidic IMER digestion enables the identification of more proteins compared to standard in-solution digestion and those proteins that are identified with both digestion methods have higher sequence coverage when digested with the IMER. CONCLUSIONS AND CLINICAL RELEVANCE The proposed reactor is well-suited for rapid and efficient protein digestion and even eight digestions can be carried out simultaneously. The PDMS chip is inexpensive and easy to fabricate, thus its application can be an attractive alternative for proteomic related research.