CEInfinite icIEF Instruments

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CEInfinite icIEF Instruments

Fast and in-depth characterisation under a variety of different conditions, including formulation and stability studies.

Our CEInfinite is a next-generation cIEF system based on a Whole Column Time-resolved Imaging Detection (WCTID). It is standard equipped with a high-sensitivity CMOS imaging sensor and a powerful deep UV LED light source that gives an increased scan rate over 100-fold compared to any other (i)cIEF system on the market.

The icIEF-Analytical simplifies protein charge analysis and provides the highest resolution separation. The icIEF fractionation and icIEF-MS modes allow you to characterise the post-translational modifications (PTMs), antibody-drug ratio (DAR), (im)purity, and chemical degradation underlying each protein charge isomer. The fractionated fractions can be used for your formulation and stability studies, such as peptide mapping, amino acid sequencing & composition, amino- & carboxy-terminal amino acids determination, disulfide bridge assessment, glycosylation & glycan characterisation, binding studies (SPR, ELISA,...), potency studies, and other biological assays

 

accelerating biologics development

Accelerating biologics development 

One icIEF workflow for your fast and in-depth product characterisation

Scan rates are over 100 times faster compared to any other imaging time-resolved cIEF systems, which enables the study of:

  • Fast dynamic processes more accurately. 
  • Empowering Taylor dispersion rapid measurement of protein hydrodynamic radius.
  • Protein interaction studies.
  • Preparative time-resolved icIEF.
  • Online (native) real-time icIEF-MS.

Analytical Profiling, Method Transfer - Existing cIEF Methods

Existing cIEF methods from most icIEF instruments can be transferred to CEInfinite Analytical instrument with no additional method development using the traditional cIEF FC-coated capillaries.

three different kinds of USP samplesThree different kinds of USP samples

The combination of the CEInfinite high sensitive detection, the use of other CEInfinite High Performance Imaging cIEF Cartridges (Patent Pending) and the AESlyte ampholytes the resolution and separation can be significantly improved.

Analytical profiling, Method Transfer - Existing CIEF Methods 

Your Advantages – the use of CEInfinite Instrument

icIEF is the industry golden standard for protein charge analysis. The CEInfinite icIEF instrument overcomes low analytical throughput and difficult method development associated with traditional cIEF, gel electrophoresis and other available icIEF instruments:

  • User-friendly interface, reliable operation control, highly efficient data processing, choose from CEInsight, Chromeleon or Empower (coming soon) Platform.
  • Fast and simplify the protein charge variant analysis.
  • Provide you consistency, repeatability and building platform methods.
  • AESlyte ampholytes and special coated cartridges for complex proteins characterisation (such as mAb, ADC, Fusion Proteins, Vaccines, mRNA, LNP and many more).
  • Easy method transfers and the utmost flexibility in method development.
  • One icIEF workflow for your fast and in-depth protein-related product characterisation (analytical profiling, fractionation, and online icIEF-MS).
  • Fast identification of PTMs, antibody-drug ratio (DAR), (im)purity and/or mass confirmation (online icIEF-MS).
  • Fractionated fractions are suitable for your formulation and stability studies, such as peptide mapping, amino acid sequencing & composition, amino- & carboxy-terminal amino acids determination, disulfide bridge assessment, glycosylation & glycan characterisation, binding studies (SPR, ELISA,...), potency studies and other biological assays.
  • High-performance CMOS imaging sensor warrants overall system compactness, energy saving and robustness
  • Full automation for fast injection and wash cycles.
  • CE and FCC certified to guarantee your safe operation

 

Whole Column Time-resolved Imaging Detection (WCTID) Technology - High Resolution Separation

The WCTID capillary electrophoresis (CE) systems have revolutionised protein charge related heterogeneity analysis with capillary isoelectric focusing (cIEF). 

  • Faster method development
  • Higher detection sensitivity
  • Better reliability
  • Higher analytical throughput

The CEInfinite icIEF is your total solution for high resolution cIEF assays. The CEInfinite icIEF instrument is standard equipped with a high sensitivity CMOS imaging sensor and powerful deep UV LED light source.

We recommend to use them in combination with the CEInfinite High Performance Imaging cIEF Cartridges (Patent Pending) and the CEInfinite AESlytes CAs, which are specially developed and synthesised for fast and reproducible high resolution cIEF assays. 

Customer review - Recent icIEF fractionation innovations with applications

Our icIEF High  Yield Fractionation and High Resolution and Reliability icIEF-MS is based on pressure mobilization. With our comprehensive icIEF pI Markers  with isoelectric points ranging from 2.85 to 11.20 and our comprehensive high and low pH carrier ampholytes, we can ensure you the maximum reproducibility in icIEF separation…

"Two principal approaches are used to fractionate charge isoform species in icIEF via the mobilization of charge isoform species: chemical mobilization and pressure mobilization. Chemical mobilization relies on the use of acetic acid or a related chemical to displace the separated zones, enabling their elution via electrophoresis. However, this approach introduces additional chemicals into the mixture, potentially leading to isoform modifications.

Furthermore, many additives used in chemical mobilization are charged and have the potential to disrupt mass spectrometry (MS) analysis through ion source contamination, such as ion suppression, which can significantly reduce sensitivity. In contrast, pressure mobilization employs pressure-driven flow to push separated isoforms to the capillary outlet. This approach ensures molecules—arise from genetic variations, alternative splicing, and post-translational modifications, leading to differences in population dynamics, cellular and tissue-specific expression, and subcellular distribution”.

Tao Bo (AES), Feng Yang (Genentech), Bo Yan (Beam Therapeutics), David A. Michels (Genentech), Tiemin Huang (AES), Janusz Pawliszyn (University of Waterloo) 2025

Read the article here

CEInfinite icIEF High Resolution - High Yield Fractionation

The charge variants are often analysed with ion exchange chromatography or a related technique capable of fractionation. However, it can take months to develop a proper method on these platforms, and there is no guarantee that they can match the resolution of icIEF. With the rise of ADCs, fusion proteins, and more, this problem will only become more pronounced.

Our Unique Solutions for you

The CEInfinite Preparative icIEF instrument provides you an unique, reliable and high resolution protein charged based separation for biologics. Utilising the custom-designed WCID cIEF-Prep cartridges (patent pending) delivers milligram-scale yields within a week.

Characterisation of individual peaks is essential for comprehending the nature of product isomers and protein degradation pathway. Therefore it is  desirable to perform peak fractionation or direct coupling to mass spectrometer for fast structure and other chemical degradation underlying characterisation of the separated protein isomer.

Proprietary cartridge - US Patent 10935519

Watch the video

Fast and High Reproducibility

Pressure mobilization is used instead of chemical mobilization to avoid instabilities related to Joule heating and pH gradient disruption. A typical single run is < 45 minutes with excellent reliability.

Avoid unstabilities. A typical single run is < 45 minutes with excellent reliability

                                                    Analytical cartridge - 100 µm                                     icIEF-Prep cartridge - 320 µm         

 

High Resolution         

A transfer capillary with a reduced diameter and specialized coatings helps minimize dispersion during transit. A reversed polarity is also possible to improve the purity of the acidic fractions.

Improve the purity of the acidic fractions.

Automation and High Throughput

The target regions for fractionation are manually selected during the first run. Each subsequent run is then automated based on this condition, thereby enabling more than 25 reproducible runs per day with minimal user intervention.

Automation and high throughput

An estimation of the fraction purity, recovery, and quantity calculated based on the a starting concentration of 2.5 mg/mL

Orthogonal Characterisation

The CEInfinite Preparative icIEF instrument can be used in combination with other analytical techniques, such as SEC-MS, to obtain a more in-depth and unique fingerprint.

High Resolution Time-resolved Imaging cIEF Fractionation – AFSC

The below work show a high resolution time-resolved imaging cIEF fractionation, fraction collection and fraction cIEF of haemoglobin control AFSC, demonstrating the speed and efficiency of the CEInfinite Preparative icIEF instrument.

High Resolution Time-resolved Imaging cIEF Fractionation – AFSC

Please contact us for further information on the materials, methods, and data analysis.

Characterizing charge heterogeneity of therapeutic antibody by icIEF-Prep and icIEF-MS

Fractionation and online mass spectrometry based on imaged capillary isoelectric focusing (icIEF) for characterising charge heterogeneity of therapeutic antibody

Learn more about the researchers at AES, how icIEF fractionation and online icIEF-MS were developed for charge variant analysis after analytical icIEF-UV profiling. The established hybrid platform was successfully utilized for in-depth charge variant characterisation of a therapeutic antibody. The fractions of a main protein component with its two major charge variants were collected for following LC-MS analysis at the intact protein level.

therapeutic antibody by icIEF-Prep and icIEF-MS

The icIEF-MS coupling was performed for rapid identification of protein heterogeneity. The whole workflow of icIEF-based fractionation and online MS detection for protein heterogeneity is straightforward, reliable, and accurate, which can provide comprehensive and cutting-edge technologies for protein drug quality control (QC) monitoring, online MS detection for rapid fingerprinting of intact proteins, and HPLC-MS in-depth characterisation of collected fractions.

Read other researchers' work:

CEInfinite icIEF-MS

The direct coupling of icIEF to a Mass Spectrometer (MS) helps you to streamline the characterisation process for biologics significantly. Characterisation of individual peaks is essential for comprehending the nature of product isomers and the protein degradation pathway. Therefore, it is desirable for direct coupling to MS for fast structure and other chemical degradation underlying characterisation of the separated protein isomer.

Or to perform peak fractionation for peptide mapping, amino acid sequencing & composition, amino- & carboxy-terminal amino acids determination, disulfide bridge assessment, glycosylation & glycan characterization, binding studies (SPR, ELISA,..), potency studies and other biological assays.

Our Unique Solutions for you

The CEInfinite icIEF-MS and Preparative icIEF instruments utilising the custom-designed WCID cIEF-MS cartridges (patent pending) for fast identification of intact charge variants. It allows you to observe real-time separation focusing (icIEF) as well as the subsequent mobilization process to isolate the charge isomer peaks.

  • High Resolution and Reliability

The pressure mobilization and transfer capillary, along with a carefully designed icIEF-MS interface, have enabled robust performance. No need to add a chemical mobilization agent to the sample matrix.

During the pressure mobilisation, an electric field keeps the samples in the separation capillary focused. The separation capillary has a larger Inner Diameter (ID), and the transfer capillary has a smaller ID. Hence, this difference greatly minimizes the remixing of separated protein isomers as they travel away from the cartridge.

High Resolution and Reliability

  • High Sensitivity - use of Critical Reagents

Use of the AESlyte carrier ampholytes and permanent MC or AD-coated cartridge for better electrospray ionization and MS signal. AESlytes are also designed for reducing the background noise that has traditionally been an issue for icIEF-MS. Formamide can be used to replace urea, which poses the same concern as MC.

aeslytes

Customer Review – Capillary icIEF-MS system and its applications (4.4)

Recent developments of icIEF technology for in-depth biopharmaceutical characterisation

“Advances in capillaries, additives, and reagents have been pivotal in overcoming the technical challenges of icIEF-MS coupling, enabling its seamless integration into high-resolution mass spectrometry workflows. The development of durable hydrophilic capillary coatings, MS-compatible additives, and innovative reagents has not only improved the sensitivity, reproducibility, and robustness of icIEF-MS but also broadened its applicability across a wide range of biopharmaceutical and analytical applications. These innovations have transformed icIEF-MS into a versatile and powerful tool for detailed protein characterisation, paving the way for its widespread adoption in both research and industrial settings”

Tao Bo (AES), Feng Yang (Genentech), Bo Yan (Beam Therapeutics), David A. Michels (Genentech), Tiemin Huang (AES), Janusz Pawliszyn (University of Waterloo) 2025

  • MS Instrument Compatibility

The platform works with various nano source MS brands, including Thermo Fisher, Waters, Bruker, Agilent and more. For high resolution and sensitivity use the Thermo Scientific Q Exactive Plus in combination with our nanospray emitter.

MS instrument compatibility

A) The transfer capillary from CEInfinite is coupled to the B) Thermo EASY-Spray Ion Source coupling using our nanospray emitter. C) This interface connects directly to the Thermo Q Exactive Plus mass spectrometer with the entire setup in approximately 5 minutes.

Thanks to the long and small transfer capillary, the CEInfinite Preparative icIEF and icIEF-MS instruments allows also to spot the separated protein elution to a MALDI target plate.

  • Seamless Method Transfer: Switching between analytical icIEF, fractionation, and icIEF-MS simply by exchanging the cartridge and limited changes to the method. As a result, this instrument provides you a highly comprehensive and flexible charge variant characterisation strategy for your biologics.
  • Makeup Solution: Acetonitrile and formic acid can be added to solution prior to MS for an improved electrospray ionization process. It is also possible to use less denaturing conditions for more sensitive molecules such as ADCs.
  • Fast identification of PTMs

The CEInfinite icIEF-MS mode enables the identification of several PTMs and chemical degradation in <45 minutes

A)icIEF profile (mAb) with 2% 8.5-10 UR AESlyte®+ 10% formamide which is mobilized towards the MS to produce a B) Total Ion Electropherogram (TIE) that displays the corresponding icIEF peaks [4]. C) Deconvoluted MS data demonstrates the identification of chemical degradation and PTMs including glycosylation, lysine residues, and deamidation.

Read other researchers' work:

Next Generation Biotherapeutics

ADCs, fusion proteins, bispecific antibodies, lipid nanoparticles (LNPs), and adeno-associated viruses (AAVs) all offer potential as the next generation of life-saving medicine. However, these modalities present unique challenges for charge variant characterisation by icIEF and IEX due to a number of factors, including increased glycosylation, conjugation, and hydrophobicity. Consequently, emerging tools for icIEF method optimization, fractionation, and direct MS coupling seem poised to play a more prominent role in biotherapeutic development.  The figure below demonstrates how our CEInfinite icIEF Solutions can provide in-depth characterization for a highly heterogeneous recombinant protein that was developed as a vaccine for SARS-CoV-2.

nextgeneration biotherapeutics icief

A collaborative study demonstrating icIEF characterisation of the recombinant protein vaccine which was extremely heterogeneous due to PTMs including N-glycans, O-glycans, deamidation, and oxidation. A) The icIEF profile and B) the corresponding TIE by MS illustrating at least 18 different charge isomers. C) The deconvoluted MS data for the first and second peaks highlighting D) extensive glycosylation as shown in the table.

We are pleased to announce our CEInfinite Chromeleon Driver integration of the CEInfinite icIEF and icIEF-MS.

It allows you to work in a well control laboratory environment and streamlining workflow while meeting 21 CFR Part 11 compliance.

CHAMALEON CEINFINITE AND IICIEF MS

Besides iCIEF-orbitrap HRMS using nano-ESI (Easy Spray), we have developed the nano-Emitter. It has been developed for high-efficient characterisation of protein drug heterogeneity, which achievement has been published in Analytical Biochemistry*

*Imaged capillary isoelectric focusing tandem high-resolution mass spectrometry using nano electrospray ionization for protein heterogeneity characterisation, Analytical Biochemistry, 2023, 680, 115312  

Have a look at our publications!

Watch Webinar - Using direct CIEF-MS and PREP-CIEF

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