Skin Tattooing As A Novel Approach For DNA Vaccine Delivery

Summary

Skin tattooing is a potent and safe way to delivery DNA vaccine intradermally. Here, a DNA plasmid encoding EGFP is delivered by tattooing to the skin of a laboratory mouse, and the expression of EGFP in the skin cells is then inspected by confocal microscopy.

Abstract

Nucleic acid-based vaccination is a topic of growing interest, especially plasmid DNA (pDNA) encoding immunologically important antigens. After the engineered pDNA is administered to the vaccines, it is transcribed and translated into immunogen proteins that can elicit responses from the immune system. Many ways of delivering DNA vaccines have been investigated; however each delivery route has its own advantages and pitfalls. Skin tattooing is a novel technique that is safe, cost-effective, and convenient. In addition, the punctures inflicted by the needle could also serve as a potent adjuvant. Here, we a) demonstrate the intradermal delivery of plasmid DNA encoding enhanced green fluorescent protein (pCX-EGFP) in a mouse model using a tattooing device and b) confirm the effective expression of EGFP in the skin cells using confocal microscopy.

Protocol

1. Plasmid DNA Purification

1. Transform the eukaryotic plasmid DNA encoding EGFP (pCX-EGFP) into DH5α E.coli competent cells. The empty pCX vector may also be used as a negative control.
2. Culture and harvest the DH5α E.coli cells and purify the pDNA according to the Qiagen EndoFree Plasmid Purification Handbook.
3. Filter pDNA solution through a 0.22 μm PVDF sterile filter, and store it at -20 °C until use.

2. Tattoo System Preparation

1. Connect the handheld unit and the control pedal to the power supply unit per the manufacturer's instructions.

Results

The expression of EGFP with an excitation peak at 488 nm and emission peak at 509 nm can be observed in mouse skin cells. From a 1.875 μg dose of DNA, containing approximately 3×10¹⁷copies of the plasmid, we typically observed 10-20 EGFP signals in the 1 cm² tattooed area. This relatively low number of transfected cells is consistent with the results of a previous study³. The EGFP expression (Figure 1) provides the evidence that EGFP plasmid was delivered into the animal'.......

Discussion

DNA vaccination is considered safer than traditional vaccination strategies as it does not require manipulation of, or expose the vaccines to, live or attenuated pathogens⁴. However, the result of DNA vaccination depends heavily on the delivery route. Skin is abundant in antigen-presenting cells, such as Langerhans Cells and dendritic cells¹, and thus an ideal site for immunization in terms of immunogenicity and ease of access^5,6. As a result, intradermal vaccination strategy is one of th.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
pCX-EGFP plasmid DNA	Clontech
STEALTH Rotary Tattoo System	Worldwide Tattoo Supply	STEALTH-L
Tattoo needles	Worldwide Tattoo Supply	1207RSB
EndoFree Plasmid Maxi Kit	Qiagen	12362
0.22 μm PVDF sterile filter	Millipore	SLGV013SL
electrical hair trimmer	Commercially available
disposable safety razors	Commercially available
Silver Sulfadiazine Cream	Watson	NDC 0591-0810-55
Ketamine HCL		NDC 0856-2012-01
Zylazine Sterile Solution		NADA 139-236