Related Experiment Video
Updated: Jul 17, 2026
10:21
Multicolor Fluorescence Detection for Droplet Microfluidics Using Optical Fibers
Published on: May 5, 2016
Instrumentation for medium-throughput two-dimensional capillary electrophoresis with laser-induced fluorescence
Cuiru Zhu1, Xinya He, James R Kraly
1Department of Chemistry, University of Washington, Seattle, Washington 98195-1700, USA.
Analytical Chemistry
|January 16, 2007
Summary
This study introduces a multiplexed two-dimensional capillary electrophoresis system for parallel analysis of five samples. The novel instrument enhances throughput for complex biological sample fingerprinting using capillary sieving electrophoresis and micellar electrokinetic capillary electrophoresis.
Area of Science:
- Analytical Chemistry
- Biochemistry
- Biotechnology
Background:
- Two-dimensional capillary electrophoresis (2D-CE) enhances separation power over 1D-CE.
- Previous 2D-CE instruments were limited to single-sample analysis, hindering throughput.
- Development of multiplexed systems is crucial for high-throughput biological sample analysis.
Purpose of the Study:
- To develop and validate a multiplexed 2D-CE instrument capable of analyzing five samples in parallel.
- To assess the performance and detection limits of the multiplexed 2D-CE system.
- To demonstrate the application of the system for generating expression fingerprints of cancer cell lines.
Main Methods:
- A novel multiplexed 2D-CE system was designed with five parallel first-dimension (capillary sieving electrophoresis - CSE) and second-dimension (micellar electrokinetic capillary chromatography - MECC) capillaries.
- Fractions were transferred between dimensions via an interface.
- Analyte detection was performed using laser-induced fluorescence in a five-capillary sheath-flow cuvette.
- The system was applied to analyze homogenates from a lung cancer (A549) cell line.
Main Results:
- The multiplexed system achieved detection limits of 940 +/- 350 yoctomoles for a labeled trypsin inhibitor, with a 3.8-fold degradation in 2D.
- An average of 131 spots (S/N > 10) were resolved per electropherogram for A549 cell line homogenates.
- Characterization of spot width and position in both CSE and MECC dimensions was performed.
- The system demonstrated a spot capacity of 300.
Conclusions:
- The developed multiplexed 2D-CE system significantly increases sample throughput for complex mixture analysis.
- The instrument provides high-resolution expression fingerprints suitable for biological sample characterization.
- This technology offers a powerful tool for proteomics and metabolomics research, particularly in cancer cell line analysis.
Related Concept Videos
01:20Capillary Electrophoresis: Instrumentation
Capillary electrophoresis instrumentation typically consists of several key components. A high-voltage power supply generates the electric field necessary for the separation by connecting to an anode (the positively charged electrode) and a cathode (the negatively charged electrode) located in buffer reservoirs at each end of the capillary tube. The system includes a sample vial, a fused silica capillary tube coated with polyimide for mechanical strength through which the sample components...
01:20Electrophoresis: Overview
Electrophoresis is a powerful analytical separation technique that relies on the differential migration of charged species when subjected to an electric field. The core strength of electrophoresis lies in its ability to separate high-molecular-weight species in complex mixtures. It has found widespread use in biochemistry, molecular biology, and analytical chemistry, allowing the separation of compounds like amino acids, nucleotides, carbohydrates, and proteins with excellent resolution.
There...
There...
01:30Capillary Electrophoresis: Applications
Capillary electrophoretic separations offer various modes, each with unique applications. These modes include capillary zone electrophoresis, capillary gel electrophoresis, capillary array electrophoresis, capillary isoelectric focusing, capillary isotachophoresis, micellar electrokinetic chromatography, and capillary electrochromatography.
Capillary zone electrophoresis (CZE) separates ionic components based on their electrophoretic mobility. It has been used to separate proteins, amino acids,...
Capillary zone electrophoresis (CZE) separates ionic components based on their electrophoretic mobility. It has been used to separate proteins, amino acids,...
01:26Inductively Coupled Plasma Atomic Emission Spectroscopy: Instrumentation
Inductively coupled plasma (ICP) is the common plasma source used in atomic emission spectroscopy (AES), a technique that detects and analyzes various elements in a sample. This method is often called inductively coupled plasma atomic emission spectroscopy (ICP-AES).
There are three main types of inductively coupled plasma atomic emission spectroscopy (ICP-AES) instruments: sequential, simultaneous multichannel, and Fourier transform instruments, with the latter being less commonly used.
There are three main types of inductively coupled plasma atomic emission spectroscopy (ICP-AES) instruments: sequential, simultaneous multichannel, and Fourier transform instruments, with the latter being less commonly used.
00:57High-Performance Liquid Chromatography: Instrumentation
High-performance liquid chromatography, or HPLC, is an analytical technique that separates liquid samples under high pressures. An HPLC instrument consists of glass bottles for storing solvents called mobile phase reservoirs. HPLC-grade solvents are used to maintain high purity, and the dissolved gases are removed using a degasser, such as a vacuum pumping system or sparging with helium. The solvents are then pumped into the analytical column using a screw-driven syringe or reciprocating pumps.
01:15High-Performance Liquid Chromatography: Types of Detectors
The role of the detectors in High-Performance Liquid Chromatography (HPLC) is to analyze the solutes as they exit from the chromatographic column. The detector recognizes the solute's property and generates corresponding electrical signals, which are converted into a readable graph of the detector's response versus elution time called a chromatogram at the computer. There are several types of HPLC detectors, each with its own advantages and limitations, depending on the analyte properties and...