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Development of an automated digestion and droplet deposition microfluidic chip for MALDI-TOF MS

  • Jeonghoon Lee
  • Harrison K. Musyimi
  • Steven A. Soper
  • Kermit K. MurrayEmail author
Article

Abstract

An automated proteolytic digestion bioreactor and droplet deposition system was constructed with a plastic microfluidic device for off-line interfacing to matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The microfluidic chips were fabricated in poly(methyl methacrylate) (PMMA), using a micromilling machine and incorporated a bioreactor, which was 100 µm wide, 100 µm deep, and possessed a 4 cm effective channel length (400 nL volume). The chip was operated by pressure-driven flow and mounted on a robotic fraction collector system. The PMMA bioreactor contained surface immobilized trypsin, which was covalently attached to the UV-modified PMMA surface using coupling reagents N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride (EDC) and hydroxysulfosuccinimide (sulfo-NHS). The digested peptides were mixed with a MALDI matrix on-chip and deposited as discrete spots on MALDI targets. The bioreactor provided efficient digestion of a test protein, cytochrome c, at a flow rate of 1 µL/min, producing a reaction time of ∼24 s to give adequate sequence coverage for protein identification. Other proteins were also evaluated using this solid-phase bioreactor. The efficiency of digestion was evaluated by monitoring the sequence coverage, which was 64%, 35%, 58%, and 47% for cytochrome c, bovine serum albumin (BSA), myoglobin, and phosphorylase b, respectively.

Keywords

PMMA Microfluidic Device Sequence Coverage Microfluidic Chip MALDI Target 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Supplementary material

13361_2011_190700964_MOESM1_ESM.doc (161 kb)
Supplementary material, approximately 165 KB.

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Copyright information

© American Society for Mass Spectrometry 2008

Authors and Affiliations

  • Jeonghoon Lee
    • 1
  • Harrison K. Musyimi
    • 2
  • Steven A. Soper
    • 1
    • 3
    • 4
  • Kermit K. Murray
    • 1
    Email author
  1. 1.Department of ChemistryLouisiana State UniversityBaton RougeUSA
  2. 2.National Institute on Aging/National Institutes of HealthBaltimoreUSA
  3. 3.Department of Mechanical EngineeringLouisiana State UniversityBaton RougeUSA
  4. 4.Center for BioModular Multi-Scale SystemsLouisiana State UniversityBaton RougeUSA

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