Name: synthesis of an in vivo mri-detectable apoptosis probe
Uploaded: 2012-07-31T12:00:00
Description: Watch this Scientific Journal Video about Synthesis of an In vivo MRI-detectable Apoptosis Probe at JoVE.com

National Institutes of Health, NHLBI (RD, PY, PS). American Heart Association (JL). Stanford University, VPUE Grant (JL).

Tilpasset fra tidligere undersøgelse 1. 1. Konjugering af Annexin V SPIO <ol><li> Oxidere SPIO partikler (Ocean Nanotech Inc., 5 mg / ml) i 1 time ved 20 ° C i mørke i opløsning med 0,15 M natriumperiodat (NalO4) (04:01 vægt: vægt-forhold) 2. </li><li> Inkubér oxiderede SPIO i 12 timer med oprenset ANX protein (1:1, vægt: vægt) i 0,15 M natriumborat (Na2 B4 O 7 10H H2O) ved 20 ° C. </li><li> Reducere blandingen i y...

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Den beskrevne konjugation fremgangsmåde til at forbinde SPIO til Annexin V udnytter sidekæde amingrupper af Annexin og carboxylgrupperne i den SPIO nanopartikel. Via specifikke oxidations-reduktions-trin, kan kovalent binding af disse forbindelser kan opnås, og den resulterende funktionaliserede nanopartikel kan isoleres. Denne metode kan generaliseres til andre proteiner og nanopartikler af interesse. De mest kritiske trin er oxidation og reduktion forhold, der udføres i de relevante te...

<table border="1"><tbody><tr><td> Navn reagens </td><td> Firma </td><td> Katalog # </td></tr><tr><td> Superparamagnetisk jernoxid </td><td> Ocean Nanotech, Inc. </td><td> ICK-40-005 </td></tr><tr><td> Doxorubicin </td><td> Sigma </td><td> D1515-10mg </td></tr><tr><td> SuperMag Separator </td><td> Ocean Nanotech, Inc. </td><td> N / A </td></tr><tr><td> Zetasizer Nano DLS maskine </td><td> Malvern, Inc </td><td> N / A </td></tr></tbody></table>

synthesis of an in vivo mri-detectable apoptosis probe

Cellulær apoptose er et fremtrædende træk ved mange sygdomme, og dette programmeret celledød forekommer typisk, før de kliniske manifestationer af sygdommen er tydelige. Et middel til at påvise apoptose i sine tidligste, reversible etaper vil give en præklinisk &#39;vindue&#39; i løbet af hvilke forebyggende eller terapeutiske kunne træffes foranstaltninger for at beskytte hjertet fra permanente skader. Vi præsenterer heri en enkel og robust metode til at konjugere human Annexin V (ANX), som ivrigt binder til celler i de tidligste og reversible stadier af apoptose, at superparamagnetisk jernoxid (SPIO) nanopartikler, der tjener som et MRI-detekterbart kontrastmiddel. Konjugeringen Fremgangsmåden begynder med en oxidation af SPIO nanopartikler, som oxideres carboxylgrupper på polysaccharidet skallen af ​​SPIO. Oprenset ANX protein tilsættes derefter i omgivelser med en natriumboratopløsning at lette kovalent interaktion ANX med SPIO i en reducerende buffer. En endelig reduktion skridt med natrium borohydride er udført for at fuldføre reduktionen, og hvorefter reaktionen standses. Ukonjugeret ANX fjernes fra blandingen ved mikrocentrifuge filtrering. Størrelsen og renheden af ​​ANX-SPIO produkt bekræftes ved dynamisk lysspredning (DLS). Denne fremgangsmåde kræver ikke tilsætning til eller ændring af polysaccharidet SPIO skallen, i modsætning til tværbundne jernoxidpartikel konjugationsfremgangsmåder eller biotin-mærkede nanopartikler. Som et resultat er denne fremgangsmåde en enkel, robust fremgangsmåde, som kan udvides til konjugering til andre proteiner af interesse.

Watch this Scientific Journal Video about Synthesis of an In vivo MRI-detectable Apoptosis Probe at JoVE.com

Synthesis of an In vivo MRI-detectable Apoptosis Probe

Tidlig detektion af apoptose kan identificere udsatte cellepopulationer i en række forskellige sygdomme. Her viser vi en metode til at forbinde en tidlig apoptose-påvisning protein (Annexin V) til et MRI-detekterbart jernoxid nanopartikler (SPIO). Denne metode kan udvides til andre proteiner af interesse at generere MRI-detekterbare molekylær billeddannelse sonder.

Syntese af en<em&gt; In vivo</em&gt; MR-påviselige Apoptose Probe

Cellular apoptosis is a prominent feature of many diseases, and this programmed cell death typically occurs before clinical manifestations of disease are ...

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Synthesis of an In vivo MRI-detectable Apoptosis Probe

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