The compact, bench-top Gelfree 8100 Fractionation Station provides specified voltage to each of eight independent channels and pauses for fraction collection according to user programming. Using the touch screen interface, the fractionation sequence is defined or selected, permitting broad mass range analyte fractionation or target protein enrichment in accordance with the users requirements. During operation, the touch screen interface provides feedback on the status and current/voltage that is applied for each channel.

Sample complexity necessitates extensive fractionation for in-depth proteomic analysis using mass spectrometry. Protein Discovery, now part of the Expedeon group, has developed a commercial system using a novel technology, termed Gel Elution Liquid Fraction Entrapment Electrophoresis (GELFREE), for molecular weight-based fractionation of intact proteins with liquid phase recovery. Eight samples are run in parallel using a programmable control module in ca. 90 minutes. The system is shown to provide robust fractionation over the mass range 3.5 kDa - 500 kDa with high loading capacity, reproducibility, and recovery.

Features

• Intact protein molecular weight fractionation, isolation, and purification
• Liquid-phase recovery without band or spot cutting
• Broad mass range fractionation up to 500 kDa
• Up to eight samples processed in parallel
• Programmable fractionation for isolating and purifying targeted proteins
• High protein recovery (>80%)
• High reproducibility (
• Sampling unbiased by hydrophobicity, pI
• High loading capacity (>5X more than a 1D gel)
• Proteins are recovered intact, for complete characterization

How it Works

Gelfree technology is "Gel-eluted Liquid Fraction Entrapment Electrophoresis.Each of the eight Gelfree 8100 Cartridge channels consists of a precision-cast gel column surrounded by pipette-accessible sample loading and fraction collection chambers. Samples are mixed with the pre-formulated Sample Buffer and pipetted into the sample loading chamber. When electric current is applied, charged molecules in the channel migrate first focus in the stacking gel and then separate according to their electrophoretic mobility or, when using SDS, molecular weight. Smaller proteins move more quickly and elute into the fractionation collection chamber, where they are trapped in a defined liquid volume. At predefined intervals, the instrument will then automatically pause for easy removal of the liquid fraction with a pipette. For collection of the next size-based fraction, the sequence is restarted and the process continued.

Performance

The Gelfree 8100 system permits effective, programmable fractionation, isolation, and purification across a broad mass range. Combined with its high load capacity and high, liquid-phase recovery , the system provides the highest sensitivity of any pre-fractionation technology. The Gelfree may be operated in fractionation mode, in which the entire sample is partitioned into 1-29 selectable MW fractions, or in targeted isolation/purification mode, allowing for extraction of analytes in one or more pre-determined mass ranges.

Broad Mass Range Fractionation

The Gelfree 8100 Fractionation System offers molecular weight-based fraction targeting over a broad mass range. Using the 12% Cartridge Kit, the system is capable of partitioning complex protein samples across the mass range 10 50 kDa. Using the 10% Cartridge Kit, the system is capable of partitioning complex protein samples across the mass range 15 100 kDa. Using the 8% Cartridge Kit, the system is capable of partitioning complex protein samples across the mass range 35 150 kDa. Using the 5% Cartridge Kit, the system is capable of partitioning complex protein samples across the mass range 75 500 kDa. Recovery across all mass ranges is > 80%.

 Intact protein molecular weight-based fractionation with liquid phase recovery

Recovery and Reproducibility

The Gelfree 8100 Fractionation System uses specialized materials to optimize protein recovery and prevent non-specific loss. In addition, each gel column used in the Gelfree system is comprised of a proprietary composite gel matrix that is robotically cast using state-of-the-art automation. Every gel tube is computer imaged to ensure compliance with strict quality control specifications and each cartridge is barcoded for tracking. This level of quality control and precision-manufacturing uniquely permits the absolute elution time reproducibility provided by the Gelfree system.

Gelfree Reproducibility of Fractionation. 200 μg yeast lysate prepared using the Gelfree 8100 Fractionation System. Fractions 1, 5, and 9 from four lanes across 2 cartridges were run on a 1D gel to demonstrate the gel-to-gel and cartridge-to-cartridge reproducibility of the system.

Programmable Methods for Targeted Isolation

The Gelfree 8100 Fractionation System is the first product to provide user-programmable molecular weight-based protein purification with liquid phase recovery. Simple adjustment of the fraction collection intervals using the touch screen interface allows isolation of analytes at a targeted molecular weight in a single fraction with near quantitative recovery.

To develop a method for targeted protein isolation using the Gelfree system, the user samples the elution window at high frequency over the expected mass range. The peak width is plotted and the fraction collection interval defined. For all subsequent runs, the optimized fraction collection interval is used to isolate and purify the target analyte in a single fraction. Fraction collection windows for a given analyte depend on the amount of total protein loaded.

Topdown proteomics community recommend GELFrEE: