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Bioengineering

Skin Tattooing As A Novel Approach For DNA Vaccine Delivery

Published: October 18th, 2012

DOI:

10.3791/50032

1Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, 2Department of Pathology, New York University School of Medicine, 3Healthcare System, Veterans Affairs New York Harbor

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.

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.

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.......

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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 th.......

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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 access5,6. As a result, intradermal vaccination strategy is one of th.......

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We would like to thank all members of the Kong Lab and Dr. Yan Deng at Microscopy Core, Office of Collaborative Science, NYUMC for their assistance and technical support. This work was supported by a pilot grant from the New York University Center for AIDS Research (CFAR, NIH grant AI027742).

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Name Company Catalog Number Comments
Name of Reagent/Material Company Catalogue 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

  1. Bins, A. D. A rapid and potent DNA vaccination strategy defined by in vivo monitoring of antigen expression. Nat Med. 11, 899-904 (2005).
  2. Pokorna, D., Rubio, I., Muller, M. DNA-vaccination via tattooing induces stronger humoral and cellular immune responses than intramuscular delivery supported by molecular adjuvants. Genet. Vaccines Ther. 6, 4 (2008).
  3. van den Berg, J. H. Optimization of intradermal vaccination by DNA tattooing in human skin. Hum. Gene Ther. 20, 181-189 (2009).
  4. Liu, M. A. DNA vaccines: a review. J. Intern. Med. 253, 402-410 (2003).
  5. Koide, Y., Nagata, T., Yoshida, A., Uchijima, M. DNA vaccines. Jpn. J. Pharmacol. 83, 167-174 (2000).
  6. Peachman, K. K., Rao, M., Alving, C. R. Immunization with DNA through the skin. Methods. 31, 232-242 (2003).
  7. Verstrepen, B. E. Improved HIV-1 specific T-cell responses by short-interval DNA tattooing as compared to intramuscular immunization in non-human primates. Vaccine. 26, 3346-3351 (2008).
  8. Potthoff, A. Immunogenicity and efficacy of intradermal tattoo immunization with adenoviral vector vaccines. Vaccine. 27, 2768-2774 (2009).
  9. Quaak, S. G. DNA tattoo vaccination: effect on plasmid purity and transfection efficiency of different topoisoforms. J. Control Release. 139, 153-159 (2009).
  10. Pokorna, D. Vaccination with human papillomavirus type 16-derived peptides using a tattoo device. Vaccine. 27, 3519-3529 (2009).

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