Name: skin tattooing as a novel approach for dna vaccine delivery
Uploaded: 2012-10-18T13:00:00
Description: Watch this Scientific Journal Video about Skin Tattooing As A Novel Approach For DNA Vaccine Delivery at JoVE.com

Vi vil gerne takke alle medlemmer af Kong Lab og Dr. Yan Deng på Mikroskopi Core, Office of Collaborative Science, NYUMC for deres bistand og teknisk support. Dette arbejde blev støttet af en pilot tilskud fra New York University Center for AIDS Research (CFAR, NIH tilskud AI027742).

1. Plasmid DNA Purification <ol><li> Omdanne eukaryotisk plasmid-DNA der koder for EGFP (pCX-EGFP) ind i DH5a E. coli kompetente celler. Den tomme pCX vektor kan også anvendes som en negativ kontrol. </li><li> Kultur og høst af DH5a E. coli-celler og oprense pDNA ifølge Qiagen EndoFree Plasmid Purification Handbook. </li><li> Filter pDNA løsning gennem et 0,22 um PVDF sterilt filter, og opbevar det ved -20 ° C indtil brug. </li></ol> 2. Tattoo System Preparation <ol><li...

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

DNA vaccination anses sikrere end de traditionelle vaccinationsstrategier da det ikke kræver manipulation af eller udsætte vaccinerne til, levende eller svækkede patogener 4. Men resultatet af DNA vaccination afhænger i høj grad om levering rute. Hud er rigelige i antigenpræsenterende celler, såsom Langerhanske celler og dendritiske celler 1, og således et ideelt sted til immunisering med hensyn til immunogenicitet og lette adgang 5,6. Som følge heraf er intradermal vaccinations...

<table border="1"><tbody><tr><td> Navn på reagens / Material </td><td> Firma </td><td> Katalognummer </td><td> Kommentarer </td></tr><tr><td> pCX-EGFP plasmid-DNA </td><td> Clontech </td><td></td><td></td></tr><tr><td> STEALTH Rotary Tattoo System </td><td> Worldwide Tattoo Supply </td><td> STEALTH-L </td><td></td></tr><tr><td> Tattoo nåle </td><td> Worldwide Tattoo Supply </td><td> 1207RSB </td><td></td></tr><tr><td> EndoFree Plasmid Maxi Kit </td><td> Qiagen </td><td> 12.362 </td><td></td></tr><tr><td> 0,22 pm PVDF sterilfilter </td><td> Millipore </td><td> SLGV013SL </td><td></td></tr><tr><td> elektrisk hår trimmer </td><td> Kommercielt tilgængelig </td><td></td><td></td></tr><tr><td> engangs barbermaskiner </td><td> Kommercielt tilgængelig </td><td></td><td></td></tr><tr><td> SølvSulfadiazin Cream </td><td> Watson </td><td> NDC 0591-0810-55 </td><td></td></tr><tr><td> Ketamin HCL </td><td></td><td> NDC 0856-2012-01 </td><td></td></tr><tr><td> Zylazine steril opløsning </td><td></td><td> NADA 139-236 </td><td></td></tr></tbody></table>

skin tattooing as a novel approach for dna vaccine delivery

Nukleinsyre-baseret vaccination er et emne af stigende interesse, især plasmid-DNA (pDNA), som koder immunologisk vigtige antigener. Efter det konstruerede pDNA administreres til vacciner er det transkriberes og translateres til immunogene proteiner, som kan fremkalde reaktioner fra immunsystemet. Mange måder at levere DNA-vacciner er blevet undersøgt, men hver leverance rute har sine egne fordele og faldgruber. Skin tatovering er en ny teknik, der er sikker, omkostningseffektiv og praktisk. Desuden kunne de punkteringer påført af nålen også tjene som en kraftigt virkende adjuvans. Her har vi a) viser den intradermale leveringsanordning af plasmid-DNA der koder for forstærket grønt fluorescerende protein (pCX-EGFP) i en musemodel ved anvendelse af en tatovering enhed og b) bekræfter den effektive ekspression af EGFP i hudceller ved hjælp af konfokal mikroskopi.

Ekspressionen af ​​EGFP med en excitations-top ved 488 nm og emission top ved 509 nm kan observeres i mus hudceller. Fra en 1,875 ug dosis af DNA, der indeholder ca 3 x 10 17 kopier af plasmidet, vi typisk set 10-20 EGFP signaler i 1 cm 2 tatoverede område. Dette relativt lave antal af transficerede celler er i overensstemmelse med resultaterne af en tidligere undersøgelse 3. EGFP ekspression (figur 1) tilvejebringer bevis for, at EGFP plasmid blev leveret ind i dy...

Watch this Scientific Journal Video about Skin Tattooing As A Novel Approach For DNA Vaccine Delivery at JoVE.com

Skin Tattooing As A Novel Approach For DNA Vaccine Delivery

Skin tatovering er en potent og sikker måde at levering DNA vaccine intradermalt. Her er et DNA-plasmid koder for EGFP leveret af tatoveringer på huden af ​​et laboratorium mus, og ekspressionen af ​​EGFP i hudceller derpå inspiceret ved konfokal mikroskopi.

Hud tatovering som en ny tilgang til DNA Vaccine Delivery

Nucleic acid-based vaccination is a topic of growing interest, especially plasmid DNA (pDNA) encoding immunologically important antigens. After the ...

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