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Biochemistry JoVE (Open Access) Citable · DOI

Microfluidic Mixers for Studying Protein Folding

DOI: 10.3791/3976-v
What you'll learn
  • ✓ Design and fabricate microfluidic mixer chips for rapid solution mixing
  • ✓ Mount, load, and operate microfluidic devices with spectroscopic detection
  • ✓ Apply UV fluorescence and FRET to analyze protein folding kinetics
  • ✓ Interpret real-time folding data from microfluidic mixer experiments
Protocol

In this work we explain the fabrication and use of a microfluidic mixer capable of mixing two solutions in ~8 μs. We also demonstrate the use of these mixers with spectroscopic detection using UV fluorescence and fluorescence resonance energy transfer (FRET).

Difficulty
advanced
Total time
~3–5 days (fabrication ~2 days; chip preparation and data collection ~1–2 days)

Steps

1
Fabricate microfluidic mixing chips

Design and manufacture microfluidic mixer devices capable of mixing two solutions in approximately 8 microseconds using standard microfabrication techniques.

▶ 01:05
2
Mount and load microfluidic chips

Install fabricated chips into a detection apparatus and introduce protein solutions and reagents for mixing experiments.

▶ 05:46
3
Collect spectroscopic data

Acquire UV fluorescence and FRET signals in real-time as mixed solutions flow through the microfluidic device.

▶ 07:35
4
Analyze protein folding kinetics

Process and interpret fluorescence data to determine protein folding rates and conformational changes following rapid mixing.

▶ 09:55
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