Capillary Zone Electrophoresis-Mass Spectrometry (CZE-MS) Technology

Capillary zone electrophoresis-mass spectrometry (CZE-MS) is an advanced analytical technique that is being applied to the characterization of biopharmaceuticals such as proteins, monoclonal antibodies, vaccines, etc. This technique combines the separation power of CZE with the sensitive detection capability of MS and can be used to identify low concentrations of proteins and their variants, which is often a requirement for biopharmaceutical analysis. CD Formulation integrates cutting-edge CZE-MSE technology into our protein characterization technology platform to provide reliable analytical support for the development and characterization of your protein/peptide drugs to determine drug purity, structural integrity and charge heterogeneity and meet regulatory requirements.

What is Capillary Zone Electrophoresis-Mass Spectrometry (CZE-MS) Technology?

CZE is an analytical chemistry technique that uses an electric field within a thin capillary filled with a buffer solution to separate ionic species based on their charge and size. CZE is characterized by high efficiency, high speed, and low sample and reagent consumption. Compared with liquid chromatography, CZE has extremely low carryover, relatively fast separation speed, and inexpensive instrumentation. CZE, coupled with mass spectrometry, provides a rapid, cost-effective analytical method to help researchers evaluate the quality, consistency, and stability of biopharmaceutical drugs during biopharmaceutical drug development.

Fig. 1 Schematic designs of the CZE-MS system. (Chen D, et al., 2023)

Capillary Zone Electrophoresis-Mass Spectrometry (CZE-MS) Principle

CZE-MS analysis is based on the separation power of electrophoresis and the sensitive detection capabilities of mass spectrometry. In CZE, an electric field is applied across a narrow capillary filled with an electrolyte solution. Charged proteins migrate through the capillary toward the electrode, and in the presence of the electric field, their migration rate is affected by their charge-to-size ratio (z/e). Smaller and more highly charged molecules migrate faster than larger or less charged molecules. As the analytes migrate, they form distinct zones based on their electrophoretic mobility. This separation occurs rapidly in the small volume of the capillary because of the high electric field strength that can be applied. The analytes exiting the capillary are ionized and analyzed by the mass spectrometer. By detecting the ions, the analytes can be identified and quantified based on their specific mass and abundance.

Our Services Related to CZE-MS

Thanks to decades of experience in supporting protein/peptide biopharmaceutical development and manufacturing using CZE-MS technology, our team of highly qualified experts offers a range of CZE-MS-related services to accelerate the implementation and success of your projects.

Our scientists will work with you to continuously optimize CZE-MS programs to support all stages of protein/peptide drug development and manufacturing - from early protein/peptide drug discovery to late-stage drug development and post-release testing under GMP.

Utilizing cutting-edge CZE-MS technology, we support the following protein/peptide characterization plans, including but not limited to:

Protein Purity Assessment

CZE-MS is an effective tool for assessing the purity of protein drug formulations, with its ability to separate and analyze multiple protein variants simultaneously. Our scientists use this technology to accelerate the development of your project by identifying any impurities and degradation products that may affect drug safety and efficacy during the development and manufacturing of proteins and peptides.

Post-Translational Modifications (PTMs) Characterization

PTMs can occur at every step of the manufacturing process. These modifications can affect the charge state of the molecule and impact the potency of the molecule. As a regulatory release assay, we utilize CZE-MS to identify these charge variants, particularly glycosylation, phosphorylation, and oxidation.

Size Exclusion and Charge Variability Assessment

CZE is able to separate proteins based on their size and charge, providing valuable insights into the heterogeneity of protein drugs. This helps understand the distribution of different protein species in a therapeutic formulation, which is critical to ensuring consistent efficacy and safety.

Protein Quantification

CZE-MS can be used to quantify protein drugs. By calibrating the response of the mass spectrometer, our scientists can determine the concentration of specific protein variants in a complex mixture, enabling precise quantification to support quality control in biopharmaceutical manufacturing.

Biosimilar Comparability Studies

CZE-MS has become an important tool for comparing the structure and function of a reference biologic with its biosimilar. This comparison is critical for regulatory approval and ensuring that biosimilars provide equivalent therapeutic benefits. Our scientists use this tool to help you obtain detailed molecular characteristics of your protein samples, including their charge heterogeneity, molecular weight distribution, and glycosylation patterns, to gain insight into the similarities and differences between the reference biologic and the biosimilar.

Our CZE-MS Equipment

In CZE-MS analysis, the analytes flowing out of CZE need to undergo a certain degree of ionization before entering the mass spectrometer for detection. To this end, our analytical laboratory is equipped with a variety of mass spectrometers with different ionization technologies to meet different analytical needs.

Available Ionization techniques for CZE analysis include:

• Electrospray ionization (ESI): This technique generates ions from the liquid phase is compatible with CZE. The electric field at the capillary outlet produces a fine mist of charged droplets containing the analyte. This technique is suitable for the analysis of more polar compounds and can achieve high sensitivity and selectivity.
• Matrix-assisted laser desorption/ionization (MALDI): Although less common in direct capillary systems, it can be used in certain configurations. This technique is more suitable for the analysis of solid samples or high molecular weight substances.

Available MS equipments for CZE analysis include:

• Quadrupole MS: Quadrupole MS can effectively select and filter ions, and combined with TOF, it can achieve high-resolution and high-sensitivity analysis.
• Time of Flight (TOF)-MS: TOF-MS can provide high-resolution mass determination, which is particularly suitable for analyzing complex mixtures.
• Ion Trap-MS: Ion Trap MS can perform multi-stage mass spectrometry analysis and has good analytical capabilities for complex samples.

Advantages of Our CZE-MS Technology

• High Resolution and Sensitivity: Due to its capillary format, CZE-MS provides excellent resolution, enabling the separation of complex protein mixtures and low-abundance protein molecules with high sensitivity.
• Rapid Analysis: CZE is inherently faster than traditional gel-based electrophoresis techniques, allowing for shorter sample turnaround and processing times.
• Minimal Sample Preparation: CZE can reduce the need for extensive sample preparation compared to other separation techniques, helping to maintain the integrity of sensitive analytes.
• Automated and Reproducible: CZE systems can be highly automated, allowing for reproducible analysis.
• Ability to Distinguish Modified Forms: CZE-MS is particularly effective in resolving different isoforms of proteins, such as phosphorylation, glycosylation, and methylation.

Custom CZE-MS Services

Why Choose Our CZE-MS Technology?

• We have a team of experts with rich experience in CZE-MS  analytical method development and validation.
• We have accumulated decades of expertise and successful project experience using CZE-MS  technology to support protein/peptide biopharmaceutical development.
• Our CZE-MS technology provides high-resolution separations and accurate mass determination, ensuring that even trace amounts of protein molecules can be reliably detected and quantified.
• We utilize the latest CZE-MS equipment and methods to provide high-quality results, ensuring your analysis is at the forefront of technology.
• We provide flexible experimental design and customized solutions.

Publication

CD Formulation aims to provide a powerful analytical tool for the separation, purification, and characterization of proteins and peptides. Please feel free to contact us if you are interested in our services. Learn how our CZE-MS technology can support the smooth implementation of your protein/peptide biopharmaceutical program.

How It Works

STEP 1

Submit a service request describing your specific research or development need.

STEP 2

We'll email you to provide your quote and confirm order details if applicable.

STEP 3

Execute the project with real-time communication, and deliver the final report promptly.