Transcutaneous Immunization Introduction for Hybridoma Generation

Transdermal immunization is one of the safe and effective immunization strategies that involves the topical application of antigens to the skin, targeting immune cells residing therein. However, the impermeability of the skin's stratum corneum makes the skin pose a strong barrier to hydrophilic macromolecules. To address this challenge, scientists have developed various methods to improve transdermal immunity.

The Role of the Skin as an Immune Organ

The skin serves as a crucial site for transcutaneous immunization, as it acts as both a physical and immune barrier due to the abundance of immunocompetent cells present within it.

Langerhans cells (LC), keratinocytes, and dermal dendritic cells (dDCs) are significant immunocompetent cells found in the skin with LC and dDCs playing a crucial role in inducing antigen-specific immune responses. Under non-inflammatory conditions, LC and dDCs are usually immature, implying their strong endocytic capacity. When external antigens enter the skin, they capture them and increase the expression of costimulatory factors, facilitating antigen presentation to T cells, CCR7 then enables the antigen-presenting cells to leave the skin and enter draining lymph nodes. Thereafter, LC and dDCs present antigens to CD4 and CD8 T cells and activate antigen-specific T and B cells.

Keratinocytes also contributed to the induction of antigen-specific immune responses by activating the innate immune system. They can efficiently convert exogenous stimuli into host homeostatic responses and express several Toll-like receptors and nucleotide-binding domain oligomerization domain-like receptors on their surface or endosomes allowing them to recognize bacterial components known as pattern-associated molecular patterns.

The mechanisms behind skin-related immunity remain incompletely understood. However, a better understanding of the cutaneous immune system can aid in further developing the role of transcutaneous immunization.

Delivery Route for Transcutaneous Immunization

Antibody production from hybridoma through transcutaneous immunization requires the selection of an appropriate delivery route. Currently, common delivery routes include electroporation, iontophoresis, sonophoresis, jet injectors, patch formulations, microneedles nanoparticles, and liposomes.

Electroporation is a method of increasing skin permeability by applying single or multiple pulses of short duration. It has been widely used to relax the cell surface, allowing the delivery of molecules into living cells. Iontophoresis therapy is a method to enhance the transport of ions or charged molecules through biological membranes by passing direct or periodic electric currents through electrolyte solutions with appropriate electrode polarity. Several groups have shown that it can facilitate the penetration of peptides or proteins, such as insulin, calcitonin, or botulinum toxin, through the stratum corneum. Sonophoresis is a method of enhancing material penetration by disrupting the stratum corneum structure with low-frequency ultrasound. Cavitation is the formation of gaseous cavities in the ultrasound-coupled medium upon exposure to ultrasound, involving either rapid growth and collapse of bubbles (transient cavitation) or slow oscillatory motion of bubbles in the ultrasound field (stable cavitation). The oscillation and collapse of cavitating bubbles perturb the lipid bilayer of the stratum corneum, thereby enhancing transport. Jet syringes are devices that use pressure to deliver substances into the skin. The patch formulation is one of the commonly used strategies for transdermal immunization, but it has many drawbacks. More convenient scientific methods, such as hydrogel patches, have been developed. Microneedle arrays contain many micron-sized needles that can create transport pathways large enough for proteins and nanoparticles, yet small enough to avoid pain. In addition, microneedle arrays can penetrate the stratum corneum barrier and deliver antigens to immunologically active cells in the skin more efficiently than other transcutaneous immunization systems

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