Name: dendra2 photoswitching through the mammary imaging window
Uploaded: 2009-06-05T00:00:00
Description: Watch this Scientific Journal Video about Dendra2 Photoswitching through the Mammary Imaging Window at JoVE.com

(; JC, JES ve JW CA100324; JvR için U54GM064346 JES CA77522, JC ve BG U54CA126511) Bu çalışma, ABD Savunma Bakanlığı&#39;nın (BG ve DK BC061403 BC075554), ABD Ulusal Sağlık Enstitüleri tarafından desteklenmiştir. D. Entenberg mikroskopi, görüntüleme kutusu ve J. Pollard (Albert Einstein Tıp Koleji) F4/80 antikor üretiminde yardım M. Rottenkolber ile yardım için teşekkür ederim.

1. Floresan tümörlerin meme yağ yastığı içine tümör hücrelerinin enjeksiyonu kullanarak Üretimi: <ol><li> % 80 confluency - 40 Dendra2 protein (sitoplazmik bir belirteci olarak) ifade eden bir hücre hattı büyütün. </li><li> PBS w / o Ca 2 yemekleri en az 3 kez durulayın + veya Mg 2 +. </li><li> 37 10 cm çanak ve inkübe başına 3 ml tripsin ekle ° C hücrelerinin çoğu ayırmak ve daha sonra düz bir yüzeye karşı çanak vurmak sallamak kadar. </li><li> Çanak ka...

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	<li>Farina, K. L., Wyckoff, J. B., Rivera, J., Lee, H., Segall, J. E., Condeelis, J. S., Jones, J. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cell+motility+of+tumor+cells+visualized+in+living+intact+primary+tumors+using+green+fluorescent+protein.">Cell motility of tumor cells visualized in living intact primary tumors using green fluorescent protein.</a> Cancer Res. 58, 2528-2532 (1998).</li><li>Naumov, G. N., Wilson, S. M., MacDonald, I., Schmidt, E. E., Morris, V. L., Groom, A. C., Hoffman, R. M., Chambers, A. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cellular+expression+of+green+fluorescent+protein,+coupled+with+high-resolution+in+vivo+videomicroscopy,+to+monitor+steps+in+tumor+metastasis.">Cellular expression of green fluorescent protein, coupled with high-resolution in vivo videomicroscopy, to monitor steps in tumor metastasis.</a> J Cell Sci. 112, 1835-1842 (1999).</li><li>Brown, E. B. <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+measurement+of+gene+expression,+angiogenesis+and+physiological+function+in+tumors+using+multiphoton+laser+scanning+microscopy.">In vivo measurement of gene expression, angiogenesis and physiological function in tumors using multiphoton laser scanning microscopy.</a> Nature Med. 7, 864-868 (2001).</li><li>Wang, W., Wyckoff, J. B., Frohlich, V. C., Oleynikov, Y., Huttelmaier, S., Zavadil, J., Cermak, L., Bottinger, E. P., Singer, R. H., White, J. G., Segall, J. E., Condeelis, J. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Single+cell+behavior+in+metastatic+primary+mammary+tumors+correlated+with+gene+expression+patterns+revealed+by+molecular+profiling.">Single cell behavior in metastatic primary mammary tumors correlated with gene expression patterns revealed by molecular profiling.</a> Cancer Res. 62, 6278-6288 (2002).</li><li>Sidani, M., Wyckoff, J., Xue, C., Segall, J. E., Condeelis, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Probing+the+microenvironment+of+mammary+tumors+using+multiphoton+microscopy.">Probing the microenvironment of mammary tumors using multiphoton microscopy.</a> J Mammary Gland Biol Neoplasia. 11, 151-163 (2006).</li><li>Wyckoff, J. B., Wang, Y., Lin, E. Y., Li, J. F., Goswami, S., Stanley, E. R., Segall, J. E., Pollard, J. W., Condeelis, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Direct+visualization+of+macrophage-assisted+tumor+cell+intravasation+in+mammary+tumors.">Direct visualization of macrophage-assisted tumor cell intravasation in mammary tumors.</a> Cancer Res. 67, 2649-2656 (2007).</li><li>Egeblad, M., Ewald, A. J., Askautrud, H. A., Truitt, M. L., Welm, B. E., Bainbridge, E., Peeters, G., Krummel, M. 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<td>PBS w/o Ca, Mg</td>
<td>&nbsp;</td>
<td>Gibco</td>
<td>14190-144</td>
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<td>Trypsin</td>
<td>&nbsp;</td>
<td>Gibco</td>
<td>25300</td>
<td>&nbsp;</td>
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<td>HBSS</td>
<td>&nbsp;</td>
<td>Gibco</td>
<td>14025</td>
<td>&nbsp;</td>
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<td>DMEM media (MDA-MB-231 cells)</td>
<td>&nbsp;</td>
<td>Gibco</td>
<td>11965</td>
<td>&nbsp;</td>
<tr>
<td>Isoflurane(Aerrane)</td>
<td>&nbsp;</td>
<td>Baxter</td>
<td># NDC 10019-773-40</td>
<td>&nbsp;</td>
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<td>SCID mouse</td>
<td>&nbsp;</td>
<td>NCI</td>
<td>N/A</td>
<td>&nbsp;</td>
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<td>&nbsp;</td>
<td>BD Falcon</td>
<td>353003</td>
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<td>Circular coverslip 8 mm</td>
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dendra2 photoswitching through the mammary imaging window

Tek hücreli çözünürlükte fare ve sıçanlarda son on yıl içinde, meme tümörlerinin intravital mikroskopi<sup&gt; 1-4</sup&gt; Mekanizmaları, göç, istila ve tümör hücrelerinin intravasation gibi metastatik davranışı önemli bilgiler sonuçlandı<sup&gt; 5, 6</sup&gt; Anjiyogenez<sup&gt; 3</sup&gt; Ve bağışıklık hücrelerinin tepki<sup&gt; 7-9</sup&gt;. Biz son zamanlarda birden çok intravital görüntüleme oturumda yaşayan farelerde görüntü ortotopik meme karsinomları için bir teknik bildirdin<sup&gt; 10</sup&gt;. Bunun için, biz Dendra2 Meme Görüntüleme Pencere (MIW) ve optimize edilmiş görüntüleme parametreleri geliştirdik<sup&gt; 11</sup&gt; Photoswitching ve görüntüleme<em&gt; In vivo</em&gt;. Burada, biz, özel bir görüntüleme kutusunu kullanarak Dendra2 etiketli tümör hücrelerinin bir tümör görüntüleme ve üstüne MIW ekleme MIW üretim için protokol tanımlar. Bu protokol görüntü metastatik tümörlerde belirgin mikroçevrelerde tümör hücrelerinin davranış için kullanılır ve kan damarları, tümör hücreleri ve ana hücreleri uzun süreli görüntüleme sağlar.

Watch this Scientific Journal Video about Dendra2 Photoswitching through the Mammary Imaging Window at JoVE.com

Dendra2 Photoswitching through the Mammary Imaging Window

Meme Görüntüleme Penceresi ile Dendra2 etiketli tümör hücrelerinin intravital photoswitching ve izleme, bize görüntü gün bir zaman ölçeği üzerinde seçilen tümör mikroçevrelerde tümör hücrelerinin metastatik davranış sağlayan bir tekniktir.

Meme Görüntüleme Penceresi yoluyla Dendra2 Photoswitching

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