Name: diagnosis of neoplasia in barrett&#8217;s esophagus using vital-dye enhanced fluorescence imaging
Uploaded: 2014-05-11T16:00:00
Description: Watch this Scientific Journal Video about Diagnosis of Neoplasia in Barrettâ€™s Esophagus using Vital-dye Enhanced Fluorescence Imaging at JoVE.com

This work is supported by the National Cancer Institute at the National Institute of Health grant R01 CA140257-01.

NOTE: Informed consent was obtained from the patients. Also, this research has been performed in compliance with all institutional, national, and international guidelines for human welfare.
1. Prepare Computer
<ol>
	<li>Turn laptop on and connect USB from the DVI2USB Capture Card.</li>
</ol>
2. Prepare Monitor
<ol>
	<li>Connect DVI cable to monitor and PinP to allow the endoscopist to view videos from these screens rather than the computer.</li>
	<l...

<ol>
	<li>Modiano, N., Gerson, L. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Barrett's+Esophagus:+Incidence,+etiology,+pathophysiology,+prevention+and+treatment.">Barrett's Esophagus: Incidence, etiology, pathophysiology, prevention and treatment.</a> Therapeutics and Clinical Risk Management. 3, 1035-1145 (2007).</li><li>Brown, L. M., Devesa, S. S., Chow, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Incidence+of+Adenocarcinoma+of+the+esophagus+among+white+Americans+by+sex,+stage,+and+age.">Incidence of Adenocarcinoma of the esophagus among white Americans by sex, stage, and age.</a> Journal of the National Cancer Institute. 100, 1184-1187 (2008).</li><li>Siegel, R., Naishadham, D., Ahmedin, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cancer+Statistics,+2012.">Cancer Statistics, 2012.</a> A Cancer Journal for Clinicians. 62, 10-29 (2012).</li><li>Vieth, M., Ell, C., Gossner, L., May, A., Stolte, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Histological+analysis+of+endoscopic+resection+specimens+from+326+patients+with+Barrett's+esophagus+and+early+neoplasia.">Histological analysis of endoscopic resection specimens from 326 patients with Barrett's esophagus and early neoplasia.</a> Endoscopy. 36, 776-781 (2004).</li><li>Pohl, H., Rosch, T., Vieth, 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=Miniprobe+confocal+laser+microscopy+for+the+detection+of+invisible+neoplasia+in+patients+with+Barrett's+oesophagus.">Miniprobe confocal laser microscopy for the detection of invisible neoplasia in patients with Barrett's oesophagus.</a> Gut. 57, 1648-1653 (2008).</li><li>Thekkek, N., 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=Modular+video+endoscopy+for+in+vivo+cross-polarized+and+vital-dye+fluorescence+imaging+of+Barrett's-associated+neoplasia.">Modular video endoscopy for in vivo cross-polarized and vital-dye fluorescence imaging of Barrett's-associated neoplasia.</a> Journal of Biomedical Optics. 18, 026007 (2013).</li><li>Roblyer, D., 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=Multispectral+optical+imaging+device+for+in+vivo+detection+of+oral+neoplasia.">Multispectral optical imaging device for in vivo detection of oral neoplasia.</a> Journal of Biomedical Optics. 13, 024019 (2008).</li><li>Egger, K., 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=Biopsy+surveillance+is+still+necessary+in+patients+with+Barrett's+oesophagus+despite+new+endoscopic+imaging+techniques.">Biopsy surveillance is still necessary in patients with Barrett's oesophagus despite new endoscopic imaging techniques.</a> Gut. 52, 18-23 (2003).</li><li>Thekkek, N., 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=Vital-dye+enhanced+fluorescence+imaging+of+GI+mucosa:+metaplasia,+neoplasia,+inflammation.">Vital-dye enhanced fluorescence imaging of GI mucosa: metaplasia, neoplasia, inflammation.</a> Gastrointestinal Endoscopy. 75, 877-887 (2012).</li><li>Muldoon, T. J., Anandasabapathy, S., Maru, D., Richards-Kortum, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-resolution+imaging+in+Barrett's+esophagus:+a+novel,+low-cost+endoscopic+microscope.">High-resolution imaging in Barrett's esophagus: a novel, low-cost endoscopic microscope.</a> Gastrointestinal Endoscopy. 68, 737-744 (2008).</li></ol>

With standard endoscopic surveillance, neoplasia in BE is often missed8 because benign metaplasia can be indistinguishable from high-grade dysplasia and adenocarcinoma. As a tool that would better enable gastroenterologists to remedy this currently unavoidable error, vital-dye enhanced fluorescence imaging highlights a tissue&#8217;s glandular morphology thereby providing a distinct feature to differentiate the tissue types. Moreover, by providing a field of view up to 2.5 cm, VFI enables endoscopists to pan o...

Over the last forty years, the incidence of esophageal adenocarcinoma (EAC) has increased significantly1,2; yet due to late diagnosis, the five-year&#160;survival rate is less than 20%3. The current standard of endoscopic surveillance in BE, the precursor to EAC, is white light endoscopy with random four-quandrant forceps&#8217; biopsies of the segment. Unfortunately, this technique often misses neoplasia, which can be flat, subtle and difficult to differentiate on standard white light imaging4. While there has been success in using confocal laser microscopy to highlight cellular features in vivo, lesions can still be missed due to the decreased field of view5. Having a &#8216;bridge&#8217; technology that can highlight areas for further confocal microendoscopic imaging would be markedly valuable.
Consequently, an enhanced red-flag imaging modality that improves the ability to target and biopsy early neoplasia in BE would be instrumental in detecting EAC at an early, curable stage and could lead to more effective treatment and subsequently improved survival rates. VFI is a novel technique that combines high-resolution epithelial imaging with exogenous topical fluorescent contrast, proflavine, to highlight glandular morphology and delineate neoplasia (high grade dysplasia and cancer) in the distal esophagus in hopes of improving the in vivo diagnosis6. Upon excitation of the proflavine, which concentrates within cell nuclei shortly after application, the fluorescent images provide spatial resolution of 50 to 100 &#956;m&#160;and a field of view up to 2.5 cm,&#160;allowing endoscopists to visualize glandular morphology. As a result, this approach enables gastroenterologists to distinguish classic Barrett&#8217;s metaplasia, which has continuous, evenly-spaced glands and an overall homogenous morphology, from BE with neoplasia, which has obliteration of the glandular architecture. Here we describe the protocol of this new technique with a multispectral endoscope, and provide representative results to demonstrate the utility of this device in depicting the morphological transformation from benign metaplasia to high-grade dysplasia and cancer.

<table border="0" cellpadding="0" cellspacing="0" width="650">
	<tbody>
		<tr>
			<td>Filter*</td>
			<td>Schott North America, Inc., Duryea, Pennsylvania</td>
			<td>Not Applicable</td>
			<td>495-nm long-pass filter</td>
		</tr>
		<tr>
			<td>Halo Cap &#8211; Medium*</td>
			<td>Barrx Medical</td>
			<td>CP-002A</td>
			<td></td>
		</tr>
		<tr>
			<td>Processor*</td>
			<td>Pentax</td>
			<td>EPK-i</td>
			<td></td>
		</tr>
		<tr>
			<td>Multispectral Digital Microscope**</td>
			<td>Not Applicable</td>
			<td>Not Applicable</td>
			<td></td>
		</tr>
		<tr>
			<td>Kimwipes</td>
			<td>Kimberly-Clark</td>
			<td>KimTech Science</td>
			<td>S-8115</td>
		</tr>
		<tr>
			<td>Cidex</td>
			<td>Advanced Sterilization Products&#160;</td>
			<td>CIDEX OPA Solution</td>
			<td></td>
		</tr>
		<tr>
			<td>Proflavine hemisulfate (0.01% w/v)</td>
			<td></td>
			<td></td>
			<td>FDA (IND 102,217)</td>
		</tr>
		<tr>
			<td>Laser Diode*</td>
			<td>Nichia Corporation</td>
			<td>Not Applicable</td>
			<td>455-nm</td>
		</tr>
		<tr>
			<td>Image Capture*</td>
			<td>Labview</td>
			<td>Not Applicable</td>
			<td></td>
		</tr>
		<tr>
			<td>Spray Catheter</td>
			<td>Olympus</td>
			<td>Not Applicable</td>
			<td></td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td>*Equipment specifics within Reference 6. **Equipment specifics within Reference 7</td>
		</tr>
	</tbody>
</table>

diagnosis of neoplasia in barrett&#8217;s esophagus using vital-dye enhanced fluorescence imaging

The ability to differentiate benign metaplasia in Barrett&#8217;s Esophagus (BE) from neoplasia in vivo remains difficult as both tissue types can be flat and indistinguishable with white light imaging alone. As a result, a modality that highlights glandular architecture would be useful to discriminate neoplasia from benign epithelium in the distal esophagus. VFI is a novel technique that uses an exogenous topical fluorescent contrast agent to delineate high grade dysplasia and cancer from benign epithelium. Specifically, the fluorescent images provide spatial resolution of 50 to 100 &#956;m&#160;and a field of view up to 2.5 cm,&#160;allowing endoscopists to visualize glandular morphology. Upon excitation, classic Barrett&#8217;s metaplasia appears as continuous, evenly-spaced glands and an overall homogenous morphology; in contrast, neoplastic tissue appears crowded with complete obliteration of the glandular framework. Here we provide an overview of the instrumentation and enumerate the protocol of this new technique. While VFI affords a gastroenterologist with the glandular architecture of suspicious tissue, cellular dysplasia cannot be resolved with this modality. As such, one cannot morphologically distinguish Barrett&#8217;s metaplasia from BE with Low-Grade Dysplasia via this imaging modality. By trading off a decrease in resolution with a greater field of view, this imaging system can be used at the very least as a red-flag imaging device to target and biopsy suspicious lesions; yet, if the accuracy measures are promising, VFI may become the standard imaging technique for the diagnosis of neoplasia (defined as either high grade dysplasia or cancer) in the distal esophagus.

Figure 1B depicts classic Barrett's Esophagus with no dysplasia surrounded on the borders by normal squamous epithelium. Beginning with the flatter squamous tissue, which is peripherally located and indicated by the blue arrows, a homogenous area of dull fluorescence is present with no glandular architecture. The green arrows indicate a circular green line surrounding the squamous tissue. This outline is artifact resulting from the cap of the endoscope. Moving to the centrally located Barrett's tissue, g...

Watch this Scientific Journal Video about Diagnosis of Neoplasia in Barrettâ€™s Esophagus using Vital-dye Enhanced Fluorescence Imaging at JoVE.com

Diagnosis of Neoplasia in Barrett&#8217;s Esophagus using Vital-dye Enhanced Fluorescence Imaging

Vital-dye enhanced fluorescence imaging (VFI) is a novel in vivo technique that combines high-resolution epithelial imaging with exogenous topical fluorescent contrast to highlight glandular morphology and delineate neoplasia (high grade dysplasia and cancer) in the distal esophagus.

The ability to differentiate benign metaplasia in Barrett&#8217;s Esophagus (BE) from neoplasia in vivo remains difficult as both tissue types can be flat ...

Research

JoVE Journal

Bioengineering

Three-dimensional Optical-resolution Photoacoustic Microscopy

Optical microscopy, providing valuable insights at the cellular and organelle levels, has been widely recognized as an enabling biomedical technology. As ...

Fluorescence Lifetime Imaging of Molecular Rotors in Living Cells

Diffusion is often an important rate-determining step in chemical reactions or biological processes and plays a role in a wide range of intracellular ...

Time-lapse Fluorescence Imaging of Arabidopsis Root Growth with Rapid Manipulation of The Root Environment Using The RootChip

The root functions as the physical anchor of the plant and is the organ responsible for uptake of water and mineral nutrients such as nitrogen, ...

High-resolution Spatiotemporal Analysis of Receptor Dynamics by Single-molecule Fluorescence Microscopy

Single-molecule microscopy is emerging as a powerful approach to analyze the behavior of signaling molecules, in particular concerning those aspect (e.g., ...

3D Orbital Tracking in a Modified Two-photon Microscope: An Application to the Tracking of Intracellular Vesicles

The objective of this video protocol is to discuss how to perform and analyze a three-dimensional fluorescent orbital particle tracking experiment using a ...

From Fast Fluorescence Imaging to Molecular Diffusion Law on Live Cell Membranes in a Commercial Microscope

It has become increasingly evident that the spatial distribution and the motion of membrane components like lipids and proteins are key factors in the ...

Fluorescence Biomembrane Force Probe: Concurrent Quantitation of Receptor-ligand Kinetics and Binding-induced Intracellular Signaling on a Single Cell

Membrane receptor-ligand interactions mediate many cellular functions. Binding kinetics and downstream signaling triggered by these molecular interactions ...

Electrotaxis Studies of Lung Cancer Cells using a Multichannel Dual-electric-field Microfluidic Chip

The behavior of directional cell migration under a direct current electric-field (dcEF) is referred to as electrotaxis. The significant role of ...

Conducting Multiple Imaging Modes with One Fluorescence Microscope

Fluorescence microscopy is a powerful tool to detect biological molecules in situ and monitor their dynamics and interactions in real-time. In addition to ...

Visualizing Adhesion Formation in Cells by Means of Advanced Spinning Disk-Total Internal Reflection Fluorescence Microscopy

In living cells, processes such as adhesion formation involve extensive structural changes in the plasma membrane and the cell interior. In order to ...