Creation of an on-chip enzyme reactor by encapsulating trypsin in sol-gel on a plastic microchip.
Trypsin-encapsulated sol-gel was fabricated in situ onto a plastic microchip to form an on-chip bioreactor that integrates tryptic digestion, separation, and detection. Trypsin-encapsulated sol-gel, which is derived from alkoxysilane, was fabricated within a sample reservoir (SR) of the chip. Fluorescently labeled ArgOEt and bradykinin were digested within the SR followed by electrophoretic separation on the same chip. The plastic microchip, which is made from poly(methyl methacrylate), generated enough electroosmotic flow that substrates and products could be satisfactorily separated. The sol-gel in the SR did not alter the separation efficiency of each peak. With the present device, the analytical time was significantly shortened compared to conventional tryptic reaction schemes. This on-chip microreactor was applicable to the digestion of protein with multiple cleavage sites and separation of digest fragments. Furthermore, the encapsulated trypsin exhibits increased stability, even after continuous use, compared with that in free solution.
Sakai-Kato
Kumiko
K
Department of Analytical Chemistry, School of Pharmaceutical Sciences, University of Shizuoka, 52-1 Yada Shizuoka, Shizuoka, 422-8526, Japan.
Kato
Masaru
M
Toyo'oka
Toshimasa
T
D004339:Drug Compounding; D004800:Enzymes, Immobilized / Q000737:chemistry*; D005782:Gels; D008832:Microchemistry / Q000295:instrumentation / Q000379:methods*; D008904:Miniaturization; D010449:Peptide Mapping / Q000295:instrumentation / Q000379:methods; D011506:Proteins / Q000032:analysis*; D014357:Trypsin / Q000737:chemistry*
D004800:Enzymes, Immobilized; D005782:Gels; D011506:Proteins; D014357:Trypsin