Name: in vivo 19f mri for cell tracking
Uploaded: 2013-11-25T21:10:00
Description: Watch this Scientific Journal Video about In vivo 19F MRI for Cell Tracking at JoVE.com

Vi vil gerne anerkende Nadine Henn og Arend Heerschap for deres værdifulde hjælp. Dette arbejde blev støttet af den nederlandske Institut for regenerativ medicin (NIRM, FES0908), EU FP7-programmet ENCITE (HEALTH-F5-2008 til 201.842) og TargetBraIn (HEALTH-F2-2012 til 279.017), et tilskud fra Volkswagen Foundation ( I/83 443), Nederlandene Organisation for Videnskabelig Forskning (VENI 700.10.409 og Vidi 917.76.363), ERC (Advanced Grant 269.019), og Radboud University Nijmegen Medical Center (AGIKO-2008-2-4).

Bemærk: Alle eksperimenter og forsøg med dyr skal udføres i overensstemmelse med de relevante etiske retningslinjer og i overensstemmelse med standard dyrepleje og humane krav. 1.. Cellemærkning (Standard Protocol med coinkubering) <ol><li> Tilføj 19F etiket 8 til umodne DC&#39;er i en koncentration på 4 mg/10 6-celler (i 2 ml medium). Sving forsigtigt for at blande. </li><li> Inkuberes cellerne med etiketten til 3 dage før modning og høst af udvikl...

<ol>
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Biotechnol. 23, 983-987 (2005).</li><li>Srinivas, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Customizable,+multi-functional+fluorocarbon+nanoparticles+for+quantitative+in+vivo+imaging+using+19F+MRI+and+optical+imaging.">Customizable, multi-functional fluorocarbon nanoparticles for quantitative in vivo imaging using 19F MRI and optical imaging.</a> Biomaterials. 31, 7070-7077 (2010).</li><li>Boehm-Sturm, P., Mengler, L., Wecker, S., Hoehn, M., Kallur, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+tracking+of+human+neural+stem+cells+with+19F+magnetic+resonance+imaging.">In vivo tracking of human neural stem cells with 19F magnetic resonance imaging.</a> PLoS One. 6, e29040 (2011).</li><li>Srinivas, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+cytometry+of+antigen-specific+t+cells+using+(19)F.">In vivo cytometry of antigen-specific t cells using (19)F.</a> MRI. 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Protokollen beskrevet her beskriver den generelle fremgangsmåde til in vivo-19F MRI cellesporing. Trods den beskrevne coinkubering metode, er der flere forskellige protokoller til mærkning af celler med et 19F middel. Imidlertid er coinkubering oftest bruges og kan optimeres yderligere fx ved tilsætning af transfektions midler 6. Den faktiske cellemærkning protokol vil afhænge af celletypen. Kun vigtige mærkningsordninger trin er beskrevet i protokollen her. Gener...

In vivo celle tracking er afgørende for optimering og overvågning af cellulære terapi 1. På grund af sin noninvasive natur, billedbehandling byder på fremragende muligheder for at overvåge celler in vivo. Magnetic Resonance Imaging (MRI) er uafhængig af ioniserende stråling og tillader en overlegen billedbehandling opløsning og iboende bløde væv kontrast. MRI-baserede celle sporing er allerede blevet anvendt klinisk til at følge dendritiske celler i melanom patienter 2. Konventionel klinisk MRI udføres på 1H kerne, til stede i mobile vand i væv. Det er også muligt at udføre MRI på andre aktive kerner, såsom 13C, 19F og 23. Na. Imidlertid har kun 19 F MRI blevet anvendt med succes til at in vivo-sporing, da det giver den højeste følsomhed efter 1. H. Fraværet af MRI-påviselige endogene 19F i væv tillader højt signal selektivitet forpåvisning af exogene 19 F kontrastmidler og tillader kvantificering af fluor direkte fra billeddataene. For en detaljeret diskussion om 19F MR, se 3-5. Et centralt problem med 19F MR er behovet for at udvikle og optimere egnede 19F celle etiketter, selv om der er udviklet flere etiketter, med en tendens til multimodale agenter 6. Protokollen vi beskriver her, er baseret på undersøgelser af vores grupper 7-9, herunder de første artikler, der er beskrevet i vivo kvantitativ 19F MRI-baserede celle sporing 10,11. Den generelle procedure for cellesporing hjælp 19F MRI er opsummeret i figur 1.. Vi beskriver en generel protokol for mærkning og billeddannelse af dendritiske celler (DCS) ved hjælp af en skræddersyet perfluorcarbon kontrastmiddel 8. De billeddannende protokollen er generelt anvendelig til forskellige celletyper, etiketter og dyremodeller. Dencelletype og dyremodel beskrevet her bør kun tages som et eksempel, og derfor har vi ikke give oplysninger om celleisolering og mærkning, men snarere fokusere på imaging-protokollen. Modifikationer vil være nødvendigt for hver etiket, celletype dyremodel og billedbehandling opsætning, og disse kan findes i litteraturen eller kan skal optimeres af forskere. Nogle almindelige ændringer er medtaget i diskussionen.

<table border="1">
		<tbody>
		 <tr>
			<td></td>
			<td></td>
			<td></td>
			<td>REAGENTS</td>
		 </tr>
		 <tr>
			<td>PBS</td>
			<td>Sigma-Aldrich</td>
			<td>MFCD00131855</td>
			<td></td>
		 </tr>
		 <tr>
			<td>TFA</td>
			<td>Sigma-Aldrich</td>
			<td>76-05-1 </td>
			<td></td>
		 </tr>
		 <tr>
			<td>Ketamine (Ketavet)</td>
			<td>Pfizer</td>
			<td>778-551</td>
			<td></td>
		 </tr>
		 <tr>
			<td>Xylazine (Rompun)</td>
			<td>Bayer</td>
			<td>QN05 cm92</td>
			<td></td>
		 </tr>
		 <tr>
			<td>Ophtosan </td>
			<td>Produlab Pharma</td>
			<td>2702</td>
			<td>eye ointment</td>
		 </tr>
		 <tr>
			<td></td>
			<td></td>
			<td></td>
			<td>Material name </td>
		 </tr>
		 <tr>
			<td>MRI scanner</td>
			<td>Bruker Biospec</td>
			<td></td>
			<td></td>
		 </tr>
		 <tr>
			<td>NMR spectrometer</td>
			<td>Bruker Biospec</td>
			<td></td>
			<td></td>
		 </tr>
		</tbody>
 </table>

in vivo 19f mri for cell tracking

In vivo 19F MRI tillader kvantitativ cellesporing uden anvendelse af ioniserende stråling. Det er en ikke-invasiv teknik, der kan anvendes til mennesker. Her beskriver vi en generel protokol for celle mærkning, billedbehandling og billedbehandling. Teknikken er anvendelig til forskellige celletyper og dyremodeller, selv om vi her at fokusere på en typisk musemodel til sporing murine immunceller. De vigtigste emner for celle mærkning er beskrevet, da disse er relevante for alle modeller. Tilsvarende er de vigtigste imaging-parametre er anført, skønt detaljerne vil variere afhængigt af MRI-system og den enkelte installation. Endelig har vi medtage et billede behandling protokol for kvantificering. Variationer for dette, og andre dele af protokollen, vurderes i diskussionen afsnit. Baseret på den detaljerede procedure der er beskrevet her, vil brugeren nødt til at tilpasse protokol for hver specifik celletype, celle etiket dyremodel, og billedbehandling setup. Bemærk, at protokol kan også tilpasses til humant brug, så længe kliniske begrænsninger overholdes.

Her viser vi de typiske resultater for en protokol der indebærer overførsel af 19 F-mærkede celler målsøgende en drænende lymfeknude. Figur 2 viser en 19F NMR-spektret af 10 6 mærkede celler under anvendelse af et TFA-reference. De billeddannende opsætning blev udført som beskrevet i protokollen (Figur 3). Til in vivo-billeddannelse, anvendte vi en reference bestående af en lukket cylinder med samme etiket anvendes til cellerne anbragt...

Watch this Scientific Journal Video about In vivo 19F MRI for Cell Tracking at JoVE.com

In vivo 19F MRI for Cell Tracking

Vi beskriver en generel protokol for in vivo-sporing ved hjælp af MRI i en musemodel med ex vivo-mærkede celler. En typisk protokol for celle mærkning, forarbejdning og kvantificering billede erhvervelse er inkluderet.

In vivo 19F MR for Cell Tracking

In vivo 19F MRI allows quantitative cell tracking without the use of ionizing radiation. It is a noninvasive technique that can be applied to humans. ...

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In vivo 19F MRI for Cell Tracking

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