Electrophoresis

Electrophoresis
Introduction
Definition
Theory
Apparatus
Loading of samples
Agarose gel Electrophoresis
Polyacrylamide gel Electrophoresis

1. 1 Clinical Laboratory Principle Electrophoresis

2. Electrophoresis Analytical Technique The most commonly used in clinical applications. Separate and analyzed Ionize analytes C harged molecules migrate in a porous supporting medium (agarose), after the sample is mixed with a buffer solution, generates an electropherogram

3. Electrophoresis Definition: “ The migration of charged solutes or particles in a liquid medium under the influence of an electrical field.”

4. Theory of Electrophoresis In an electrophoresis system, chemical take on electrical charge and becomes ionized. Move toward electrode N egative E lectrode :Cathode P ositive Electrode : Anode Positive ions(cations ) migrate toward the Cathode Negative ions ( anions) migrate toward the Anode

5. Theory of Electrophoresis Cathode Anode

6. Electrophoresis Apparatus Buffer Tanks Electrodes (Anode/Cathode) Electrophoretic support for separation Wick Tank Cover Power Supply

7. Electrophoresis Apparatus

8. Electrophoresis Agarose G el E lectrophoresis (AGE ) Polyacrylamide Gels Electrophoresis (PAGE)

9. Agarose G el E lectrophoresis (AGE ) Smaller DNA fragments have migration rates in agarose that are inversely proportional to the logs of their molecular weights This relationship decreases as their fragment size increases.

10. Loading of PCR Amplified Product in Agarose Gel

11. Direction Vertical Horizontal

12. Agarose Gel Electrophoresis (AGE)

13. Agarose Gel Electrophoresis (AGE)

14. Visualization of PCR Amplified DNA Products

15. Polyacrylamide Gels ( PAGE) Polyacrylamide is a polymer Prepared by heating acrylamide with a variety of catalysts, with or without cross-linking agents. Polyacrylamide gel is Thermostable Transparent Durable Chemically I nert (uncharged)

16. Polyacrylamide Gels C apable of resolving DNA molecules that differ by as little as 2% in length (1 bp in 50 bp ). Accommodates a larger amount of sample (up to 10 fig) in a single sample slot (well) DNA recovered from a polyacrylamide gel is extremely pure, containing no inhibitors . Polyacrylamide is most useful for mixtures of smaller DNA fragments and resolves fragments < 1 kbp

17. Apparatus for Polyacrylamide Gels (PAGE)