Development of an enzymatic reactor applying spontaneously adsorbed trypsin on the surface of a PDMS microfluidic device
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Herein, a microfluidic device (MD) containing immobilized trypsin for rapid and efficient proteolysis was described. Trypsin was immobilized via non-specific protein adsorption onto the hydrophobic poly(dimethylsiloxane) (PDMS) channel wall of the MD. Peptide mapping of bovine serum albumin (BSA) samples was carried out to estimate the stability of trypsin adsorbed on PDMS surface. Peptide maps of BSA samples were obtained by capillary zone electrophoresis (CZE), the RSD% for migration times were under 1%. Several proteins (hemoglobin, myoglobin, lysozyme, and BSA) in a wide molecular size range (15–70 kDa) were digested efficiently with ∼50 s contact time. The number of separated peaks correlated well with the expected number of peptides formed in the complete tryptic digestion of the proteins. Peptide mass fingerprinting of BSA and human serum was carried out. Trypsin retained its activity for 2 h; within this period, the MD can be used for multiple digestions. The main properties of this device are simple channel pattern, simple immobilization procedure, regenerability, and disposability; all these features make this MD one of the simplest yet applicable enzymatic microreactors.
KeywordsImmobilization Trypsin Digestion Peptide mapping Enzyme reactor Microfluidic device
The research was supported by the EU and co-financed by the European Regional Development Fund under the project GINOP-2.3.2-15-2016-00008, GINOP-2.3.3-15-2016-00004, and NTP-EFÖ-P-15-0003 project. The authors also acknowledge the financial support provided to this project by the National Research, Development and Innovation Office, Hungary (K111932). We would like to thank Prof. Dr. Csaba Hegedűs for measurement possibilities in the Biomaterials Research Lab on the Faculty of Dentistry.
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Conflict of interest
The authors declare that they have no conflict of interest.
- 46.Pawliszyn J. Solid phase microextraction: theory and practice. Wiley-VCH; 1997.Google Scholar