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Related Concept Videos

Protein Dynamics in Living Cells01:19

Protein Dynamics in Living Cells

Different fluorescence-based techniques are used to study the protein dynamics in living cells. These techniques include FRAP, FRET, and PET.
Fluorescent recovery after photobleaching (FRAP) is a fluorescent-protein-based detection technique used to quantify protein movement rates within the cell. This method exposes a small portion of the cell to an intense laser beam. The laser beam causes permanent photobleaching of the fluorophore-tagged proteins in the exposed region. As the bleached...

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Related Experiment Video

Updated: May 8, 2026

A Microfluidic Chip for the Versatile Chemical Analysis of Single Cells
15:41

A Microfluidic Chip for the Versatile Chemical Analysis of Single Cells

Published on: October 15, 2013

Probing enzymatic activity inside single cells.

Jessica Olofsson1, Shijun Xu, Gavin D M Jeffries

  • 1Department of Chemical and Biological Engineering, Chalmers University of Technology , Kemivägen 10, SE-412 96 Gothenburg, Sweden.

Analytical Chemistry
|September 6, 2013
PubMed
Summary
This summary is machine-generated.

This study introduces a rapid microfluidic method to measure enzyme activity in single cells. The novel approach allows for quick, quantitative analysis of intracellular enzyme kinetics and inhibition at the cellular level.

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Area of Science:

  • Biochemistry
  • Cell Biology
  • Microfluidics

Background:

  • Enzyme activity is crucial for cellular function.
  • Measuring enzyme kinetics at the single-cell level presents significant challenges.
  • Existing methods often require large cell populations or are time-consuming.

Purpose of the Study:

  • To develop a novel, rapid method for quantifying enzymatic activity within individual suspended cells.
  • To characterize alkaline phosphatase activity and inhibition in single NG108-15 cells.
  • To establish dose-response and dose-inhibition curves at the single-cell level.

Main Methods:

  • Utilized a steady-state microfluidic device for controlled cell permeabilization with digitonin.
  • Exposed mildly permeabilized single cells to varying concentrations of fluorescein diphosphate (FDP) and levamisole.
  • Monitored enzymatic activity using fluorescence detection.

Main Results:

  • Successfully generated quantitative estimates of intracellular enzyme activity in single cells.
  • Determined apparent Michaelis constant (Km) and inhibition constant (Ki) for alkaline phosphatase.
  • Achieved rapid measurements, with activity monitored within 40 seconds and dose-inhibition curves within 150 seconds.

Conclusions:

  • This microfluidic approach provides a fast and effective new methodology for single-cell enzyme characterization.
  • The technique enables the construction of kinetic and inhibition profiles at the individual cell level.
  • Offers potential for high-throughput screening and detailed cellular mechanistic studies.