We would like to acknowledge Nadine Henn and Arend Heerschap for their valuable assistance. This work was supported by the Netherlands Institute of Regenerative Medicine (NIRM, FES0908), the EU FP7 program ENCITE (HEALTH-F5-2008-201842) and TargetBraIn (HEALTH-F2-2012-279017), a grant from the Volkswagen Foundation (I/83 443), the Netherlands Organization for Scientific Research (VENI 700.10.409 and Vidi 917.76.363), ERC (Advanced Grant 269019), and Radboud University Nijmegen Medical Centre (AGIKO-2008-2-4).

Note: All experiments and procedures involving animals must be carried out in accordance with relevant ethical guidelines and conform with standard animal care and humane requirements.
1. Cell Labeling (Standard Protocol with Coincubation)
<ol>
	<li>Add 19F label8 to immature DCs at a concentration of 4 mg/106 cells (in 2 ml medium). Swirl gently to mix.</li>
	<li>Incubate the cells with the label for 3 days before maturing and harvesting the DCs.</li>
	<li>C...

<ol>
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W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fluorine+(19F)+MRS+and+MRI+in+biomedicine.">Fluorine (19F) MRS and MRI in biomedicine.</a> NMR Biomed. 24, 114-129 (2011).</li><li>Stoll, G., Basse-Lusebrink, T., Weise, G., Jakob, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Visualization+of+inflammation+using+19F-magnetic+resonance+imaging+and+perfluorocarbons.">Visualization of inflammation using 19F-magnetic resonance imaging and perfluorocarbons.</a> Wires Nanomed. Nanobiol. 4, 438-447 (2012).</li><li>Srinivas, M., Boehm-Sturm, P., Figdor, C. G., de Vries, I. 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The authors have no conflicts of interests to disclose.

The protocol outlined here describes the general procedure for in vivo 19F MRI cell tracking. Despite the described co-incubation method, there are several different protocols for labeling cells with a 19F agent. However, the co-incubation is most often used and can be further optimized e.g. by addition of transfection agents6. The actual cell labeling protocol will depend on the cell type. Only key labeling steps are described in the protocol here. Generally, the label ...

In vivo cell tracking is essential for the optimization and monitoring of cellular therapeutics1. Due to its noninvasive nature, imaging offers excellent opportunities to monitor cells in vivo. Magnetic Resonance Imaging (MRI) is independent of ionizing radiation and allows a superior imaging resolution and intrinsic soft tissue contrast. MRI-based cell tracking has already been used clinically to follow dendritic cells in melanoma patients2. Conventional clinical MRI is carried out on the 1H nucleus, present in mobile water in tissues. It is also possible to carry out MRI on other active nuclei, such as 13C, 19F and 23Na. However, only 19F MRI has been applied successfully to in vivo cell tracking as it offers the highest sensitivity after 1H. The absence of MRI-detectable endogenous 19F in tissues permits high signal selectivity for the detection of exogenous 19F contrast agents and allows quantification of fluorine concentration directly from the image data. For a detailed discussion on 19F MRI, see3-5. A key issue with 19F MRI is the need to develop and optimize suitable 19F cell labels, although several labels have been developed, with a trend towards multimodal agents6.
The protocol we describe here is based on studies by our groups7-9, including the first articles that described in vivo quantitative 19F MRI-based cell tracking10,11. The general procedure of cell tracking using 19F MRI is summarized in Figure 1. We describe a general protocol for labeling and imaging of dendritic cells (DCs) using a custom-made perfluorocarbon contrast agent8. The imaging protocol is generally applicable to different cell types, labels and animal models. The cell type and animal model described here should only be taken as an example, and thus we do not provide details on the cell isolation and labeling, but rather focus on the imaging protocol. Modifications will be necessary for each label, cell type, animal model, and imaging setup, and these can be found in the literature or may need to be optimized by the researchers. Some common modifications are included in the discussion.

<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 allows quantitative cell tracking without the use of ionizing radiation. It is a noninvasive technique that can be applied to humans. Here, we describe a general protocol for cell labeling, imaging, and image processing. The technique is applicable to various cell types and animal models, although here we focus on a typical mouse model for tracking murine immune cells. The most important issues for cell labeling are described, as these are relevant to all models. Similarly, key imaging parameters are listed, although the details will vary depending on the MRI system and the individual setup. Finally, we include an image processing protocol for quantification. Variations for this, and other parts of the protocol, are assessed in the Discussion section. Based on the detailed procedure described here, the user will need to adapt the protocol for each specific cell type, cell label, animal model, and imaging setup. Note that the protocol can also be adapted for human use, as long as clinical restrictions are met.

Here we show the typical results for a protocol involving the transfer of 19F-labeled cells homing to a draining lymph node. Figure 2 shows a 19F NMR spectrum of 106 labeled cells using a TFA reference. The imaging setup was carried out as described in the protocol (Figure 3). For the in vivo imaging, we used a reference consisting of a sealed cylinder of the same label used for the cells placed between the feet of the mouse. A representative pro...

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

In vivo 19F MRI for Cell Tracking

We describe a general protocol for in vivo cell tracking using MRI in a mouse model with ex vivo labeled cells. A typical protocol for cell labeling, image acquisition processing and quantification is included.

In vivo 19F MRI 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. ...

Research

JoVE Journal

Medicine

14.6K visualizações.  Radboud University Medical Center.  Nós descrevemos um protocolo geral para no rastreamento de células in vivo utilizando ressonância magnética em um modelo de rato com ex vivo células marcadas. Um protocolo típico para a marcação de células, o processamento de aquisição de imagem e a quantificação é incluído. 