Paper Electrophoresis

Introduction

• Paper electrophoresis is one of the earliest forms of electrophoretic techniques.

• It employs cellulose paper as the medium through which molecules migrate under an electric field.

Paper Electrophoresis

Principle

• Molecules dissolve in a buffer solution and migrate through the porous structure of the paper based on their charge and size when an electric field is applied.

Methodology

Preparation of the Paper:

• Cut the filter paper strip to the desired size and soak it in a buffer solution to maintain stable pH conditions.

Sample Application:

• Apply a small, precise spot of the sample at a designated point on the paper to minimize spreading and maintain resolution.

Electrophoresis Chamber Setup:

• Position the paper in a chamber containing a buffer solution.

• Ensure both ends of the paper strip are in contact with the buffer for proper conductivity.

• Place electrodes (anode and cathode) at opposite ends of the chamber.

Application of Electric Field:

• Connect the chamber to a power supply to establish an electric field across the paper.

• Charged molecules in the sample migrate toward the electrode with the opposite charge based on their charge-to-mass ratio.

Detection and Analysis:

• After electrophoresis, remove the paper and detect separated molecules using staining or other visualization methods.

• Analyze the migration distances to identify and characterize the sample components.

Types of Paper Electrophoresis

1) High Voltage Paper Electrophoresis:

• Voltage: Above 2000V.

• Duration: Faster separation due to higher voltage.

• Resolution: Better resolution with sharper bands.

• Applications: Ideal for rapid separations and smaller molecules.

2) Low Voltage Paper Electrophoresis:

• Voltage: Below 300V.

• Duration: Slower separation due to lower voltage.

• Resolution: Gentler process but may result in less sharp bands.

• Applications: Suitable for larger molecules or sensitive samples.

Advantages

1. Simplicity: Easy to perform with minimal equipment.

2. Cost-Effective: Inexpensive materials and setup.

3. Portability: Suitable for field applications and educational purposes.

4. Visualization: Clear band formation facilitates qualitative analysis.

5. Versatility: Can separate a variety of small to moderately sized molecules.

Disadvantages

1. Low Resolution: Limited ability to separate complex or closely related molecules.

2. Quantitative Limitations: Primarily qualitative with limited accuracy for quantification.

3. Reproducibility Issues: Variability due to manual handling and environmental factors.

4. Sensitivity: Lower sensitivity compared to more advanced electrophoretic techniques.

5. Sample Capacity: Limited sample loading capacity, restricting the analysis of large quantities.

Applications of Paper Electrophoresis

1. Amino Acid Separation: Differentiating amino acids based on charge and size.

2. Clinical Diagnostics: Analyzing blood proteins and other biomarkers.

3. Pharmaceutical Analysis: Quality control of drugs and active ingredients.

4. Forensic Science: Identification of substances in forensic samples.

5. Educational Demonstrations: Teaching fundamental electrophoretic principles in laboratories.