Experion™ Automated Electrophoresis System

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Overview

The Experion™ automated electrophoresis system employs LabChip microfluidic technology to automate protein and nucleic acid electrophoresis. It integrates separation, detection, and data analysis within a single platform. Using much smaller sample and reagent quantities than standard analysis method, the Experion system accomplishes analysis in a single 30–40 minute, automated step. This section provides an overview of the technology behind the Experion system and describes the system and its capabilities.

Related Topics: Automated Electrophoresis and Analysis.

Applications of Experion System

The Experion system is used for the following applications:

  • Protein analysis — quantitation and size determination (sizing) of protein sample useful for quality control, protein purity and stability analysis, protocol optimization, and evaluation of recombinant protein expression
  • RNA analysis — evaluation of RNA integrity and concentration. The system automatically generates the RNA quality indicator (RQI), a quantitative integrity assessment, in addition to visual electropherogram data, ribosomal RNA ratios, and concentration data. This makes Experion RNA assays useful upstream of applications such as quantitative real-time PCR (qPCR), microarray analysis, sequencing, and reverse-transcription qPCR (RT-qPCR)
  • DNA analysis — quantitation and sizing of restriction digests, amplified DNA, microsatellites, and amplified fragment length polymorphisms (AFLPs)

The Experion system. The system includes the following components: 1) automated electrophoresis station, 2) priming station, 3) vortex station used for nucleic acid analysis only, 4) system operation and data analysis tools (software), and 5) analysis kits, which include the (a) chips and (b) reagents for protein (Pro260 kit), standard sensitivity RNA (StdSens kit), high-sensitivity RNA (HighSens kit), and DNA (DNA 1K and 12K kits) analyses.

System Components

Electrophoresis Station
The Experion automated electrophoresis station houses all the electrical, optical, and hardware components necessary to perform the separation, staining, destaining, detection, and imaging of samples in a single step. The electrophoresis station is equipped with an internal universal power supply that operates at 100–240 VAC, 50–60 Hz. It connects to a computer (PC) running the Experion software.

Experion electrophoresis station.

Priming Station
The Experion priming station is used to prepare the microfluidic chips for sample analysis. The priming station applies pressure to the priming well of the chip and fills the network of microchannels with the gel-stain solution (a mixture of Experion gel and stain).

Experion priming station.

Vortex Station
The Experion vortex station ensures complete mixing of RNA or DNA samples and loading buffers in the chip wells. The vortex adapter securely holds the chip during the 1 min vortex cycle. The vortex station is not used for protein analysis.

Experion vortex station.

Microfluidic Chips
At the center of the Experion system is the microfluidic chip in which sample separation occurs. Each chip contains a series of plastic wells bonded over a small glass plate. The glass plate is etched with a network of microchannels that connects with the base of the plastic wells. The microchannels are primed, or filled with a mixture of gel matrix and stain, before the samples are applied to the wells. Once a prepared chip is placed in the electrophoresis station and the lid closed, 16 electrode pins contact the solutions in each well. The electrophoresis station directs the samples through the microchannels by controlling the voltages and currents that are applied. Different chips are used for each application.

Experion microfluidic chips.

Experion microfluidic chips.

Analysis Kits: Microfluidic Chips and Reagents
Experion analysis kits contain the microfluidic chips and reagents required to perform protein, RNA, or DNA separation and analysis (see table below).

  • Experion Pro260 analysis kit — used to resolve and quantitate 10–260 kD proteins with sensitivity comparable to that of colloidal Coomassie Blue gel staining. The kit includes the Experion Pro260 ladder, a modified version of Bio-Rad's Precision Plus recombinant proteins of 10–260 kD for accurate sizing of samples analyzed on a chip. Each chip can separate up to 10 samples per analysis
  • Experion RNA StdSens and RNA HighSens analysis kits — offer rapid single-step assessments of RNA quality and quantity and the ability to analyze 1–12 samples in approximately 30 min. Sample total RNA or mRNA concentrations in nanogram (StdSens) or picogram (HighSens) amounts can be analyzed
  • Experion dsDNA 1K and DNA 12K analysis kit — used for single-step sizing and quantitation of dsDNA fragments in up to 11 samples
  Pro260 RNA HighSens RNA StdSens DNA 1K DNA 12K
# Samples 1–10 1–11 1–12 1–11 1–11
Sample volume 4 µl 1 µl 1 µl 1 µl 1 µl
Concentration range 5–2,000 ng/µl 100–5,000 pg/µl 5–500 ng/µl 0.1–50 ng/µl 0.1–50 ng/µl
Separation range 10–260 kD 15–1,500 bp 50–17,000 bp
Sensitivity 2.5 ng/µl
(carbonic anhydrase
in 1x PBS)
100 pg 5 ng 0.1 ng 0.1 ng

 

Experion Software
Experion software includes the controls required to operate the electrophoresis station and the data analysis tools needed to assess the results. During a run, the software provides the ability to monitor run progress or review previous sample results. For more information about the use of Experion software, refer to the Experion analysis kit manuals in the Documents tab.

Two optional software kits assess system performance and aid in troubleshooting:

  • The Experion validation kit includes automated instrument qualification (IQ) and operational qualification (OQ) protocols that test the critical functions of the system and verify and validate specific system functionality; it is useful for troubleshooting
  • Experion Security Edition software includes the IQ/OQ protocols of the validation kit in addition to tools for compliance with U.S. FDA 21CFR Part 11 regulations

Experion Software. Experion software provides a user-friendly interface for protein, RNA, or DNA analysis. For total RNA analysis, the software automatically provides such elements as A, an electropherogram for visualizing RNA profile; B, a results table showing an RQI value with convenient color coding rankings along with a run summary; and C, a tree-view browser for organizing runs with D, virtual gel.

Overview of Technology

Automated electrophoresis systems use microfluidic chip based technology to integrate electrophoretic separation, fluorescence detection, and data analysis of various biomolecules such as proteins and nucleic acids within a single platform. The Experion analysis chip utilizes LabChip technology developed by Caliper Life Sciences, Inc. The microfluidic chip is a powerful, miniaturized device — much like a tiny laboratory — that combines the functionality of a number of larger benchtop analytical instruments. Within each chip, a series of microchannels connects the sample wells to a separation channel and buffer wells. A set of electrodes in the electrophoresis station applies a voltage across the microchannels, causing charged molecules in the samples to migrate into and through the separation channel. Samples are run sequentially, with a sufficient lag between them to prevent cross-contamination. For separation, the microchannels are filled with a proprietary gel-stain solution (GS) that acts as a sieving matrix.

  • Sample RNA fragments migrate through the separation channel at a rate based on their size and charge. RNA samples are heat-denatured to minimize the effects of shape or structure on migration. During separation, the fluorescent dye intercalates between the bases of the RNA fragments
  • Under denaturing conditions in the presence of lithium dodecyl sulfate (LDS), sample proteins migrate through the separation channel at a rate based on their size. During separation, the fluorescent dye associates with the LDS micelles coating the proteins and with free micelles. An important difference between the Experion system and traditional sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) lies in how sample fragments are detected: in SDS-PAGE, samples are generally stained in the gel once separation is completed, while in the Experion system proteins are stained with a fluorescent dye during separation. Furthermore, non-reducing conditions can be run within a single protein chip run, without reducing agent contamination to other wells within a chip
  • Sample dsDNA fragments migrate through the separation channel at a rate based on their size and charge. During separation, the fluorescent dye intercalates between the bases of the fragment

Finally, the fragments with the fluorescent dye bound (directly bound to RNA or DNA or bound to LDS micelles coating the proteins) are detected as they pass a laser and photodiode detector (laser-induced fluorescence). Data are automatically collected, saved, and presented in several formats (electropherogram — fluorescence vs. time trace, virtual gel, and data tables) by the Experion software.

Microfluidic technology enables much lower consumption of sample and reagent quantities and speeds up the run and analysis time compared to standard analysis methods. Microfluidic automated electrophoresis systems such as the Experion system reduce hands-on time and enable completion of protein or nucleic acid electrophoresis and analysis of up to 12 samples in a single 30–40 min assay (analysis of a single sample is accomplished in as little as 90 sec). The Experion system can be used in applications such as purity and yield assessment in protein purification and protocol optimization workflows, RNA integrity assessments before RT-qPCR or next generation sequencing, or separation and detection of DNA fragments from PCR reactions or RFLP analysis. Although samples cannot be recovered from the separations, sample consumption is minimal, hence enabling preservation of more of the original sample for other applications.

Protein, RNA, or DNA analysis with the Experion system is achieved with various analysis kits, which supply the microfluidic chips, reagents, and instructions required to separate and analyze 10–260 kD proteins under denaturing conditions, total or mRNA, and double stranded DNA.

Data are displayed by the Experion software and show visual plots such as electropherograms and virtual gels, and linear regression curves, information such as size, concentration, % contribution of a particular separated species against the total sample, RNA quality number. The types of key data and abilities are available include the following:

Data or function

Protein Assay

RNA assays

DNA assays

Electropherogram (plot of fluorescence vs. time)

X

X

X

Virtual gel (densitometric gel like image)

X

X

X

Overlaying electropherograms of samples within a run or across different runs

X

X

X

Manual integration of peaks*

X

X

X

Data summary table

X

X

X

Size information

kD

nt

bp

Concentration

ng/µl range**

ng/µl or pg/µl range

ng/µl range

%Total sample (% contribution of each peak detected in sample)

X

 

X

RQI (RNA quality indicator) for objective RNA integrity assessment

 

X

 

Calibration curve and use of user chosen internal standard

X

 

 

 

*  Available for Ladder lane only for RNA assays
**Pro260 assay sensitivity is comparable to colloidal Coomassie (Brilliant) Blue staining of SDS-PAGE gels

Experion Workflow

Sample analysis involves the following steps:

  • Preparing the chip (priming, loading, and vortexing) — Priming fills the microchannels of the microfluidic chip with the gel-stain solution, which contains both the sieving matrix and fluorescent dye. Samples prepared in Experion sample buffer are then loaded in the sample wells. The chip is vortexed in RNA and DNA analyses
  • Running the chip — The chip is inserted into the electrophoresis station, and as the electrophoresis station lid is closed, electrodes come in contact with the solution in the wells. The voltage applied to the sample wells causes the charged molecules to migrate into the separation channel. In the separation channel, the gel-stain solution causes the molecules to move at different rates, depending on their size. During separation, the fluorescent dye, which is specific to the analysis being performed, associates with the lithium dodecyl sulfate (LDS) coating the proteins (protein analyses) or intercalates between the bases of RNA (RNA analyses) or DNA (DNA analyses)
  • Detecting the molecules — As the molecules migrate toward the end of the separation channel, a laser excites the dye, causing it to fluoresce if it is bound to the protein-LDS micelle complexes or RNA/DNA fragments. A photodiode detects the fluorescence, and Experion software plots the fluorescence intensity as a function of time to produce an electropherogram. A virtual gel image is generated from the electropherogram data
  • Data analysis — Once separation is complete, the software subtracts background noise, identifies and integrates peaks, and assigns their sizes and concentrations; the results of data analysis are automatically tabulated and presented in a table. Experion software displays all three forms of data simultaneously (see figure below)

Related Content

Experion System Training
The four part training videos presented here provides information on the Experion automated electrophoresis system, helps you refine your chip loading techniques, understand software features and shows an example of a succesful RNA run.
 
Using the Experion Software for Protein Analysis
Using the Experion Software for Protein Analysis
This webinar by Field Application Specialist Sean Taylor presents the advanced features of the Experion software for accurately calculating protein concentrations by using customizable internal standard and standard curve calibration samples.
You will be taken to our collaborator (Blackboad Collaborate) site, from where you can download the entire presentation.
 
Literature
Number Description Download
10000975 Instruction Manual, Experion Pro260 Analysis Kit, Rev C Click to download
10000976 Instruction Manual, Experion RNA StdSens and HighSens Analysis Kits, Rev C Click to download
10001312 Instruction Manual, Experion Automated Electrophoresis System, Rev C Click to download
10004490 Quick Guide, Experion Pro260 Analysis Kit, Rev C Click to download
10004491 Quick Guide, Experion RNA HighSens Analysis Kit, Rev B Click to download
10004492 Quick Guide, Experion RNA StdSens Analysis Kit, Rev B Click to download
10006451 Quick Guide, Experion DNA 1K Analysis Kit, Rev B Click to download
10006453 Instruction Manual, Experion DNA 1K and DNA 12K Analysis Kits, Rev B Click to download
10006455 Quick Guide, Experion DNA 12K Analysis Kit, Rev B Click to download
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