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 ønsker å takke alle medlemmer av Kong Lab og Dr. Yan Deng på Mikroskopi Core, Office of Collaborative Science, NYUMC for deres assistanse og teknisk støtte. Dette arbeidet ble støttet av en pilot stipend fra New York University Center for AIDS Research (CFAR, NIH stipend AI027742).

1. Plasmid DNA Rensing <ol><li> Transformere eukaryote plasmid DNA-koding EGFP (PCX-EGFP) i DH5α E.coli kompetente celler. Den tomme PCX vektoren kan også bli brukt som en negativ kontroll. </li><li> Kultur og høste DH5α E.coli celler og rense pDNA henhold til Qiagen EndoFree Plasmid Rensing Handbook. </li><li> Filter pDNA løsningen gjennom et 0,22 um PVDF sterilt filter, og lagre det ved -20 ° C inntil bruk. </li></ol> 2. Tattoo System Forberedelse <ol><li> Koble den ...

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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-vaksinering anses tryggere enn tradisjonelle vaksinasjonsstrategier som det ikke krever manipulasjon av, eller utsett vaksiner til, live eller dempes patogener 4. Imidlertid avhenger resultatet av DNA-vaksinering tungt på levering ruten. Skin er rik på antigen-presentasjon celler, for eksempel Langerhans celler og dendrittiske celler 1, og dermed et ideelt område for immunisering i form av immunogenitet og enkel tilgang 5,6. Som et resultat, er intradermal vaksinasjon strategi en ...

<table border="1"><tbody><tr><td> Navn på reagens / Material </td><td> Selskapet </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 forsyning </td><td> STEALTH-L </td><td></td></tr><tr><td> Tatovering nåler </td><td> Worldwide Tattoo forsyning </td><td> 1207RSB </td><td></td></tr><tr><td> EndoFree Plasmid Maxi Kit </td><td> Qiagen </td><td> 12362 </td><td></td></tr><tr><td> 0,22 mikrometer PVDF sterilt filter </td><td> Millipore </td><td> SLGV013SL </td><td></td></tr><tr><td> elektrisk hår trimmer </td><td> Kommersielt tilgjengelig </td><td></td><td></td></tr><tr><td> disponibel barberbladene </td><td> Kommersielt tilgjengelig </td><td></td><td></td></tr><tr><td> SølvSulfadiazine 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 Sterile Solution </td><td></td><td> NADA 139-236 </td><td></td></tr></tbody></table>

skin tattooing as a novel approach for dna vaccine delivery

Nukleinsyre-basert vaksinasjon er et tema av økende interesse, spesielt plasmid DNA (pDNA) koding immunologisk viktige antigener. Etter den utviklet pDNA administreres til vaksiner, er det transkriberes og oversatt til immunogen proteiner som kan fremkalle reaksjoner fra immunsystemet. Mange måter å levere DNA-vaksiner er undersøkt, men hver leveranse rute har sine egne fordeler og fallgruver. Skin tatovering er en ny metode som er trygg, kostnadseffektiv og praktisk. I tillegg kunne de punkteringer påført av nålen også tjene som et potent adjuvans. Her har vi a) demonstrere intradermal levering av plasmid DNA-koding forbedret grønt fluorescerende protein (PCX-EGFP) i en mus modell med en tatovering enhet og b) bekrefte effektiv uttrykk for EGFP i hudcellene ved hjelp konfokalmikroskopi.

Uttrykk for EGFP med en eksitasjon topp på 488 nm og utslipp topp på 509 nm kan observeres i mus hudceller. Fra en 1.875 mikrogram dose av DNA, som inneholder ca 3 × 10 17 eksemplarer av plasmidet, vi vanligvis observert 10-20 EGFP signaler i 1 cm 2 tatovert området. Dette relativt lave antall transfekterte celler er konsistent med resultatene av en tidligere undersøkelse 3. Den EGFP uttrykk (figur 1) gir bevis for at EGFP plasmid ble levert inn i dyrets hud celler ...

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 trygg måte å levering DNA-vaksine intradermalt. Her er en DNA-plasmid som koder for EGFP levert av tatovering til huden av et laboratorium mus, og ekspresjon av EGFP i hudcellene deretter inspisert av konfokal mikroskopi.

Skin tatovering som en ny tilnærming for DNA vaksine levering

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

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