Name: ex vivo culture of patient tissue &amp; examination of gene delivery
Uploaded: 2010-12-20T18:00:00
Description: Watch this Scientific Journal Video about Ex Vivo Culture of Patient Tissue & Examination of Gene Delivery at JoVE.com

This work was funded through the Cork South Infirmary Victoria University Hospital Breast fund, the Irish Cancer Society (CRI07TAN) and the Cork Cancer Research Centre. Replication incompetent recombinant Adenovirus 5 particles under the transcriptional control of the CMV promoter was a kind gift from Prof. Andrew Baker, University of Glasgow.

Preparation of Media and Reagents
<ol>
 <li>Antibiotic Solution 
 Phosphate buffered saline (PBS) with 200 U/ml penicillin, 200 &mu;g/ml streptomycin, 5 &mu;g/ml fungizone</li>
 <li>Collection and Treatment medium 
 Dulbecco's modified eagle medium (DMEM) with 200 U/ml penicillin, 200 &mu;g/ml streptomycin, 5 &mu;g/ml fungizone</li>
 <li>Culture medium 
 Collection medium containing 10% heat-inactivated fetal bovine serum</li>
</ol>
<p class=...

<ol>
	<li>van de Bovenkamp, M., Groothuis, G. M., Draaisma, A. L., Merema, M. T., Bezuijen, J. I., van Gils, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Precision-cut+liver+slices+as+a+new+model+to+study+toxicity-induced+hepatic+stellate+cell+activation+in+a+physiologic+milieu.">Precision-cut liver slices as a new model to study toxicity-induced hepatic stellate cell activation in a physiologic milieu.</a> Toxicol Sci. 85, 632-638 (2005).</li><li>Speirs, V., Green, A. R., Walton, D. S., Kerin, M. J., Fox, J. N., Carleton, P. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Short-term+primary+culture+of+epithelial+cells+derived+from+human+breast+tumours.">Short-term primary culture of epithelial cells derived from human breast tumours.</a> Br J Cancer. 78, 1421-1429 (1998).</li><li>Kirby, T. O., Rivera, A., Rein, D., Wang, M., Ulasov, I., Breidenbach, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+novel+ex+vivo+model+system+for+evaluation+of+conditionally+replicative+adenoviruses+therapeutic+efficacy+and+toxicity.">A novel ex vivo model system for evaluation of conditionally replicative adenoviruses therapeutic efficacy and toxicity.</a> Clin Cancer Res. 10, 8697-8703 (2004).</li><li>Josserand, V., Texier-Nogues, I., Huber, P., Favrot, M. C., Coll, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Non-invasive+in+vivo+optical+imaging+of+the+lacZ+and+luc+gene+expression+in+mice.">Non-invasive in vivo optical imaging of the lacZ and luc gene expression in mice.</a> Gene Ther. 14, 1587-1593 (2007).</li></ol>

We describe an ex vivo patient tissue culture method and bioluminescent detection system for the assessment of gene delivery within non-fixed human tissue. The method offers a simple and reproducible way of culturing tissue slices. It has significant potential as it allows the study of gene delivery into intact human tissue, the analysis of gene delivery into a variety of human tissue including malignant tissue and can provide important information concerning the effects of the high degree of variability between...

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		<th>Material Name</th>
		<th>Type</th>
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		<th>Catalogue Number</th>
		<th>Comment</th>
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<td>DMEM</td>
<td>&nbsp;</td>
<td>Sigma-Aldrich</td>
<td>D6429</td>
<td>&nbsp;</td>
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<td>PBS</td>
<td>&nbsp;</td>
<td>Sigma-Aldrich</td>
<td>D8537</td>
<td>&nbsp;</td>
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<td>Firefly Luciferin</td>
<td>&nbsp;</td>
<td>Biosynth, Switzerland</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
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<td>Dermabond</td>
<td>&nbsp;</td>
<td>Johnson &amp; Johnson</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
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<td>Vibrotome</td>
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ex vivo culture of patient tissue &amp; examination of gene delivery

This video describes the use of patient tissue as an ex vivo model for the study of gene delivery. Fresh patient tissue obtained at the time of surgery is sliced and maintained in culture. The ex vivo model system allows for the physical delivery of genes into intact patient tissue and gene expression is analysed by bioluminescence imaging using the IVIS detection system. The bioluminescent detection system demonstrates rapid and accurate quantification of gene expression within individual slices without the need for tissue sacrifice. This slice tissue culture system may be used in a variety of tissue types including normal and malignant tissue and allows us to study the effects of the heterogeneous nature of intact tissue and the high degree of variability between individual patients. This model system could be used in certain situations as an alternative to animal models and as a complementary preclinical mode prior to entering clinical trial.

Watch this Scientific Journal Video about Ex Vivo Culture of Patient Tissue & Examination of Gene Delivery at JoVE.com

Ex Vivo Culture of Patient Tissue & Examination of Gene Delivery

This article describes the culture of patient tissue slices for gene delivery studies and subsequent analysis of gene expression using IVIS bioluminescence imaging.

Ex Vivo Culture of Patient Tissue &amp; Examination of Gene Delivery

This video describes the use of patient tissue as an ex vivo model for the study of gene delivery.  Fresh patient tissue obtained at the time of surgery ...

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