Electrophoresis

Author: Mary Ellen Koenn, MS, MLS(ASCP)
Reviewer: Leslie Lovett, MS, MLS(ASCP) and Joshua J. Cannon, MS, MLS(ASCP)CMSHCM
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Continuing Education Credits

P.A.C.E.® Contact Hours: 1.5 hour(s)

Florida Board of Clinical Laboratory Science CE Credit Hours

Florida Board of Clinical Laboratory Science CE - General (Clinical Chemistry/UA/Toxicology): 1.5 hour(s)

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This course discusses the many types and applications of electrophoresis in the clinical laboratory. Commonly used terms are defined and procedures are described, including explanations of various electrophoretic patterns. A discussion on newer molecular diagnostic techniques that incorporate electrophoresis conclude this in depth look at electrophoresis.

Objectives

  • Explain the principle of electrophoresis and compare the factors that determine the mobility and rate of migration in electrophoresis.
  • Describe the role of the buffer in electrophoresis, types of support mediums, and the kinds of specimens used in electrophoresis.
  • Compare and contrast the following types of electrophoresis: routine electrophoresis, polyacrylamide gel electrophoresis, high resolution electrophoresis, capillary electrophoresis, isoelectric focusing, two-dimensional electrophoresis, pulsed field electrophoresis, and immunochemical electrophoresis.
  • Define each of the following terms: amphoteric, ampholyte(s), isoelectric point, sodium dodecyl sulfate, cathode, and anode.
  • Name and describe the stains used in electrophoresis.
  • Evaluate causes of irregular, distorted or atypical electrophoresis bands, along with correct sample application and proper care and use of buffers and support mediums.
  • Explain electroendosmosis and wick flow as encountered in electrophoresis and assess their effect on separated bands.
  • Discuss the theory behind and uses of densitometry.
  • Discuss the theory behind and uses of densitometry.
  • Discuss the theory behind and uses of densitometry.

Course Outline

  • Basic Principals of Electrophoresis
    • Introduction
    • Principle of Electrophoresis
    • Amphoteric
    • Isoelectric Point (pI)
    • Mobility or Rate of Migration
    • Rate of Migration
    • Which statement is correct for a protein with an isoelectric point (pI) of 7.0?
    • The pI of a protein is 9.2. This protein is placed in an electrical field where a buffer sets the pH at 10.0. Select the correct statement regarding the electrophoretic migration of this protein.
    • Which one of the following will slow down the migration of solutes in electrophoresis?
  • Electrophoresis and Buffers
    • Role of Buffers
    • Buffers and pH
    • True or False: Proteins in a buffer with the pH set at 8.6 will become anions and move to the positively charged electrode.
  • Specimens for Electrophoresis
    • Specimens
    • After reviewing the information on specimen samples for electrophoresis, select the correct statement.
  • Electrophoresis and Support Media
    • Types of Support Media
    • Agarose Gel
    • Polyacrylamide Gels
    • There are several different types of media that can be used in electrophoresis. Most methods today use a gel (cellulose acetate, agarose, or polyacrylamide). Which one of the following statements is true regarding these gels?
  • Electrophoresis Instrumentation
    • Electrophoresis Equipment
    • Automation
    • True or False: Automated electrophoresis systems only include automated reagent addition, electrophoresis of samples, staining of separated fragments, and detection of separated bands.
  • Types of Electrophoresis
    • Types of Electrophoresis
    • Routine Electrophoresis
    • High Resolution Electrophoresis (HRE)
    • Polyacrylamide Electrophoresis (PAGE)
    • Denaturing Polyacrylamide Gels
    • Capillary Electrophoresis (CE)
    • CE Advantages and Applications
    • Isoelectric Focusing (IEF)
    • IEF Advantages and Applications
    • Immunofixation Electrophoresis
    • Pulsed Field Electrophoresis
    • Two-Dimensional Electrophoresis
    • Two-Dimensional Electrophoresis Advantages and Applications
    • Summary: Types of Electrophoresis
    • In isoelectric focusing, the basis of separation of solutes is different than the other types of electrophoresis. Which statement below correctly describes this feature of isoelectric focusing?
    • True or False: Sodium dodecyl sulfate is added to polyacrylamide gels to denature the proteins in the sample and enhance their separation.
  • Visualization and Detection
    • Visualization and Detection Methods
    • Stains and Dyes
    • Densitometry
    • Labeled Probes
    • Which statements below are correct descriptions of visualization and detection methods used in electrophoresis?
  • Technical Considerations and Electrophoresis Troubleshooting
    • Technical Considerations and Electrophoresis Troubleshooting Topics
    • Sample Application
    • Buffers
    • Support Media
    • Electroendosmosis
    • Wick Flow
    • Troubleshooting Irregular, Distorted, or Atypical Bands
    • Which statements below are associated with electroendosmosis?
    • True or False: Wick flow is caused by the movement of the buffer ions into the medium when moisture loss due to heat generation occurs.
  • Resurgence of Electrophoresis
    • Resurgence of Electrophoresis
    • Electrophoresis and Molecular Diagnostics
    • Blotting Techniques
    • Uses of CE in Molecular Diagnostics
  • References
    • References

Additional Information

Level of Instruction: Intermediate
Intended Audience: Medical laboratory scientists, medical laboratory technicians, laboratory supervisors, and laboratory managers. This course is also appropriate for MLS and MLT students and pathology residents.
Author Information: Mary Ellen Koenn, MS, MLS(ASCP), is an associate professor emeritus at West Virginia University School of Medicine, Medical Laboratory Science Program. She has been a medical technologist and educator, having been a laboratory supervisor and manager and holding several teaching positions. She has authored numerous articles for laboratory publications and textbook chapters and frequently presents at laboratory seminars and workshops. Ms. Koenn holds a Master of Science degree in Medical Technology.
The author has no conflict of interest to disclose.
Reviewer Information:
Leslie Lovett, MS, MLS(ASCP), is the Clinical Education Coordinator of the Medical Laboratory Technology Program and a professor at Pierpont Community and Technical College in West Virginia. She holds an MS in Medical Technology with classes toward a PhD in Pharmacology and Toxicology.
Joshua J. Cannon, MS, MLS(ASCP)CMSHCM received his Bachelor of Science and Master of Science in Medical Laboratory Science from Thomas Jefferson University in Philadelphia, PA. He holds Medical Laboratory Scientist and Specialist in Hematology certifications through the ASCP Board of Certification. He was a professor at Thomas Jefferson University for seven years before transitioning into his current role as Education Developer at MediaLab by Vastian. His areas of expertise and professional passions include clinical hematology and interprofessional education.
Course Description: This course discusses the many types and applications of electrophoresis in the clinical laboratory. Commonly used terms are defined, and procedures are described, explaining various electrophoretic patterns. A discussion on molecular diagnostic techniques that incorporate electrophoresis concludes this in-depth look at electrophoresis.