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Capillary Electrophoresis Presentation
 

Capillary Electrophoresis Presentation

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Introduction to Beckman Coulter CE

Introduction to Beckman Coulter CE

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    Capillary Electrophoresis Presentation Capillary Electrophoresis Presentation Presentation Transcript

    • P/ACE MDQ Capillary Electrophoresis System
    • Capillary Electrophoresis: A Simple Technique
      • I n 1989, Beckman Instruments introduced the first fully automated capillary electrophoresis instrument (P/ACE™ 2000) to the scientific community. At that time, CE demonstrated exceptional resolving of selected compounds, but the new technology lacked a track record of applications. The subsequent application of automated CE to real-world separation problems has propelled the advancement of this technology to the robust, dedicated analyzers in use today.
    • Separation Technology
      • Capillary Zone Electrophoresis (CZE), also known as free-solution CE (FSCE), is the simplest form of CE. The separation mechanism is based on differences in the charge-to-mass ratio of the analytes.
      • Capillary Gel Electrophoresis (CGE) is the adaptation of traditional gel electrophoresis into the capillary using polymers in solution to create a molecular sieve also known as replaceable physical gel.
      • Capillary Isoelectric Focusing (CIEF) allows amphoteric molecules, such as proteins, to be separated by electrophoresis in a pH gradient generated between the cathode and anode.
      • Isotachophoresis (ITP) is a focusing technique based on the migration of the sample components between leading and terminating electrolytes. Solutes having mobilities intermediate to those of the leading and terminating electrolytes stack into sharp, focused zones. Although it is used as a mode of separation, transient ITP has been used primarily as a sample concentration technique.
      • Electrokinetic Chromatography (EKC) is a family of electrophoresis techniques named after electrokinetic phenomena, which include electroosmosis, electrophoresis and chromatography.
      • Micellar Electrokinetic Capillary Chromatography (MECC OR MEKC) is a mode of electrokinetic chromatography in which surfactants are added to the buffer solution at concentrations that form micelles. The separation principle of MEKC is based on a differential partition between the micelle and the solvent.
      • Micro Emulsion Electrokinetic Chromatography (MEEKC) is a CE technique in which solutes partition with moving oil droplets in buffer. The microemulsion droplets are usually formed by sonicating immicible heptane or octane with water. SDS is added at relatively high concentrations to stabilize the emulsion.
      • Non-Aqueous Capillary Electrophoresis (NACE) involves the separation of analytes in a medium composed of organic solvents. The viscosity and dielectric constants of organic solvents affect both sample ion mobility and the level of electroosmotic flow.
      • Capillary Electrochromatography (CEC) is a hybrid separation method that couples the high separation efficiency of CZE with HPLC and uses an electric field rather than hydraulic pressure to propel the mobile phase through a packed bed.
    • Applications of CE
      • Genetic analysis
      • Analysis of pharmaceuticals
      • Pharmaceuticals with chiral centers (enantiomers)
      • Counter-ion analysis in drug discovery
      • Protein characterization
      • Carbohydrate analysis for the determination of post translational modifications
    • Pharmaceutical Analysis
      • The highly polar nature of pharmaceuticals containing basic amine functional groups makes the use of chromatography quite complex. Ion pairing reagents and stringent column regeneration is often necessary to reduce nonspecific ionic interactions that occur with reverse-phase chromatography. With CE, these highly functional amines are favored and may be exploited to provide extraordinary resolution. The most common and simplified format of operation is to use bare-fused silica capillaries at a well defined acidic pH. Under these conditions the capillary surface is essentially non-reactive while an analyte's amine functional groups are maximally ionized, rendering a simplified robust assay for the analysis of basic drugs. The P/ACE MDQ, coupled with photo-diode array detection, is being used for this application, and is being applied effectively in the pharmaceutical industry for the analysis of basic drugs, pharmokinetic profiling, bioavailability determinations, plasma protein binding studies and drug activity level determination.
      • One of the challenges of drug discovery is in developing analytical methods for the pharmacokinetic (PK) profiling of new drug candidates. Important to this process is the development of rapid, generic methods that allow the screening of large numbers of compounds isolated from complex sample matrices. CE is being used routinely to quantify drugs in blood plasma, as well as in brain, kidney and heart tissue. The high efficiency and the lack of interference from the matrix make CE a fast and easy analytical tool for PK screening. A derivative of this application is also being used for screening drugs of toxicological interest, enabled by combining an analytes spectral signature with its electrophoretic mobility to provide highly reproducible identification.
    • Process Flow
    • CE Principle Electrophoretic Migration Electroosmotic Flow
    • Electropherogram Detector Output Migration Time Cation (+) Neutral Anion (-)
    • Anode + Cathode -
    • Advantages of CE
      • Simple with selective separation mechanism
      • High resolving power
      • Fast analysis time
      • Nanoliter sample injection
      • Minimal reagent consumption
      • High degree of system automation
      • Interface on-line with other instruments
    • Instrumentation Autosampler Module Fluid Delivery Module XZY Robotics Module Detector Modules Capillary Containment Module Capillary Temperature Control Module