Abstract: Transfecting blood cells with nucleic acids is a valuable technique for both research and clinical applications. Electroporation has emerged as an effective method for introducing nucleic acids into blood cells by applying short, high-voltage electric pulses. This review provides an overview of electroporation-based methods for transfecting blood cells, focusing on optimization strategies to enhance transfection efficiency and minimize cell damage. Additionally, we discuss the applications of electroporation-mediated transfection in both research and clinical settings, highlighting its potential in gene therapy, immunotherapy, and other therapeutic interventions.

Introduction: Transfection of blood cells plays a crucial role in advancing our understanding of cellular processes, developing novel therapeutics, and personalized medicine. Electroporation, also known as electrotransfer, is a technique that employs short, high-intensity electric pulses to create transient pores in cell membranes, enabling efficient uptake of nucleic acids. This review aims to provide insights into the use of electroporation for transfecting blood cells, focusing on optimization strategies and applications in research and clinical settings.

Electroporation-Based Transfection Methods:

1. Bulk Electroporation: a. Suspension Electroporation: Blood cells are suspended in a conductive buffer and subjected to electric pulses in cuvettes or specialized electroporation chambers. b. Microfluidic Electroporation: Blood cells are introduced into microchannels, and electric pulses are applied to facilitate transfection.
2. Single-Cell Electroporation: a. Microinjection Electroporation: Individual blood cells are microinjected with nucleic acids, followed by the application of electric pulses to enhance transfection efficiency. b. Dielectrophoresis-Assisted Electroporation: Blood cells are trapped and aligned in an electric field, allowing precise delivery of nucleic acids into selected cells.

Optimization Strategies:

1. Pulse Parameters: Optimization of pulse duration, amplitude, and number of pulses is crucial for achieving high transfection efficiency while minimizing cell damage.
2. Buffer Composition: The choice of electroporation buffer, including pH, osmolarity, and ionic strength, can significantly impact transfection efficiency.
3. Nucleic Acid Delivery: Optimization of nucleic acid concentration, purity, and delivery method ensures effective transfection and gene expression.
4. Cell Handling: Factors such as cell density, viability, and temperature during electroporation influence transfection outcomes.

Applications in Research and Clinical Settings:

1. Gene Therapy: Electroporation enables the delivery of therapeutic genes into blood cells for the treatment of genetic disorders, cancer, and other diseases.
2. Immunotherapy: Transfecting immune cells, such as T cells or dendritic cells, allows the expression of specific receptors or antigens, enhancing their therapeutic potential in cancer immunotherapy.
3. Drug Discovery: Electroporation-mediated transfection facilitates the screening of potential drug targets, evaluation of drug efficacy, and investigation of cellular pathways.
4. Hematopoietic Stem Cell Engineering: Transfection of hematopoietic stem cells enables genetic modification for gene therapy approaches or enhancing their engraftment potential in transplantation.

Conclusion: Electroporation-based transfection methods offer versatile and efficient means to introduce nucleic acids into blood cells for various research and clinical applications. Optimization strategies for pulse parameters, buffer composition, nucleic acid delivery, and cell handling are critical to achieving high transfection efficiency. Electroporation holds great promise in gene therapy, immunotherapy, drug discovery, and hematopoietic stem cell engineering, fostering advancements in personalized medicine and therapeutic interventions.