Tutorial & Science Café Vendor Technical Workshops

Tutorial

Capillary Iso-Electric Focusing (CIEF) – Prime Methodology for Protein Characterization
Presented by Dr. Gerard Rozing, ROZING.COM Consulting, Karlsruhe, Germany
Tutorial Description: Since the inception of iso-electric focusing (IEF) as a high-resolution electrophoretic technique for the separation of amphoteric substances by Tiselius and Vestenberg, it has become an indispensable method for the separation of protein mixtures. Initially, IEF was executed on a flat surface coated with a gel mixed with ampholytes to establish the pH gradient. The sample is applied to the center of the plate. This so-called slab gel IEF became more sophisticated by using flat surfaces with an immobilized pH gradient (IPG). With the introduction of instruments for Capillary Electrophoresis, the technique was adapted to allow execution in-the capillary and subsequent, online optical detection (capillary iso-electric focusing, CIEF). This was followed by dedicated whole-column-image-detection electrophoresis systems (iCIEF). With the commercialization of CIEF, the methodology has evolved into a recipe-driven, black-box approach. The users strictly follow a protocol provided by the system supplier. Therefore, the basic understanding of the separation mechanism on IEF has faded. Diagnosis and troubleshooting when “it does not work” become an insurmountable hurdle, and the costly help of the system supplier is called. Therefore, there is a continuing need and effort for teaching the basic principles of iso-electric focusing, especially when applied in capillaries (CIEF). In this tutorial lecture, the author will emphasize on: 

a.          Basics of iso-electric focusing and in capillary execution. 

b.          Method development of CIEF. 

c.          Instrumentation for CIEF and iCIEF. 

d.          Tools for capillary iso-electric focusing method development. 

e.          Preparative CIEF. 

f.          Coupling of CIEF with Mass Spectrometric Detection. 

In the spirit of the MSB symposia series, the author will fill 2/3 of the allocated time on presentation leaving 1/3 for Q&A. 

Science Café Vendor Technical Workshops

 

  • Bruker (gold sponsor) Workshop on "Latest Applications of 4D-Proteomics using Trapped Ion Mobility on the timsTOF Pro 2; and 4D Proteomics – Dissecting the 3D Structure of Proteins through Ion Mobility Enhanced Crosslinking Mass Spectrometry"
  • SCIEX (gold sponsor) Workshop on "Comprehensive, 15-min Charge Variant Analysis of Biotherapeutics with a Microfluidic Chip-Based Integrated iCIEF-MS System"
  • VICI (gold sponsor) Workshop on "Microscale Separations at Nanoscale"
  • Agilent (silver sponsor) Workshop on "Advancing Denaturing and Native Top-down Proteomics Analysis using CE-MS"
  • Phenomenex (silver sponsor) Workshop on "Combining the Power of a Core-Shell Particles and Advanced Stationary Phase Selectivity to Improve Micro and Nano Flow Separations"
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