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Highly Sensitive and Quantitative Detection of Proteins and Their Isoforms by Capillary Isoelectric Focusing Method
07:58

Highly Sensitive and Quantitative Detection of Proteins and Their Isoforms by Capillary Isoelectric Focusing Method

Published on: September 19, 2018

Application of imaging capillary IEF for characterization and quantitative analysis of recombinant protein charge

Zoran Sosic1, Damian Houde, Andy Blum

  • 1Analytical Development, Biogen Idec, Cambridge, MA 02142, USA. zoran.sosic@biogenidec.com

Electrophoresis
|November 20, 2008
PubMed

Insights

Imaging capillary isoelectric focusing (icIEF) offers a superior method for analyzing charge heterogeneity in recombinant proteins and monoclonal antibodies. This technique provides faster development, quicker analysis, and higher throughput compared to traditional methods.

Area of Science:

  • Biochemistry
  • Analytical Chemistry
  • Protein Chemistry

Background:

  • Charge heterogeneity is a critical quality attribute for recombinant proteins and monoclonal antibodies.
  • Traditional methods like gel electrophoresis and ion-exchange chromatography (IEC) have limitations in speed and throughput for charge heterogeneity analysis.

Purpose of the Study:

  • To evaluate the application of imaging capillary isoelectric focusing (icIEF) for charge heterogeneity analysis of recombinant proteins and monoclonal antibodies.
  • To demonstrate the advantages of icIEF over traditional methods.
  • To qualify icIEF for quality control and stability monitoring.

Main Methods:

  • Imaging capillary isoelectric focusing (icIEF) was employed for charge heterogeneity assessment.
  • Correlation between icIEF and ion-exchange chromatography (IEC) was established.
  • Electrospray ionization mass spectrometry (ESI-MS) was used for identification of charged variants.

Main Results:

  • icIEF demonstrated advantages in speed, throughput, and method development time compared to gel and IEC.
  • A strong correlation was observed between icIEF-detected isoforms and IEC-determined charge heterogeneity.
  • Intermediate precision for pI determination was ≤0.2% RSD, with relative peak area reproducibility within acceptable limits for main and acidic species.

Conclusions:

  • icIEF is a powerful and efficient technique for quantitative analysis of charge heterogeneity in recombinant proteins and monoclonal antibodies.
  • The method is suitable for quality control environments, enabling robust monitoring of protein stability and identity.
  • icIEF offers fast method development, short analysis times, and high sample throughput, making it advantageous for biopharmaceutical development.

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