Name: a novel technique for generating and observing chemiluminescence in a biological setting
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Description: Watch this Scientific Journal Video about A Novel Technique for Generating and Observing Chemiluminescence in a Biological Setting at JoVE.com

The authors thank Prof. Jan Grimm and Mr. Travis Shaffer for their helpful discussions and Mr. David Gregory for editing the manuscript. Technical services provided by the MSK Animal Imaging Core Facility, supported in part by NIH Cancer Center Support Grant P30CA008748-48, are gratefully acknowledged. The authors thank the NIH (K25 EB016673 and R21 CA191679, T.R. and 4R00CA178205-02, B.M.Z.), the MSK Center for Molecular Imaging and Nanotechnology (T.R.), the Tow Foundation (B.C.), and the National Science Foundation Integrative Graduate Education and Research Traineeship (IGERT 0965983 at Hunter College for B.C. and T.M.S.) for their generous support. The research reported in this publication was supported by funding from the King Abdullah University of Science and Technology.

Ethiek statement: Alle in vivo dierexperimenten beschreven werden uitgevoerd volgens een goedgekeurd protocol en in het kader van de ethische richtlijnen van het Memorial Sloan Kettering Cancer Center (MSK) Institutional Animal Care en gebruik Comite (IACUC). 1. Bouw van een vernevelen Device <ol><li> Bevestig hout deel A (12,5 x 2,5 x 1,8 cm 3) rechtop in het midden van deel B (12,7 x 10,7 x 1,8 cm 3) met twee schroeven (4 x 25 mm 2). Attach hout ...

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Hier hebben wij voorgesteld een technologie die in staat is optisch afbakenen weefsel via de emissie van fotonen door een chemiluminescente reporter is. In tegenstelling tot andere, meer gevestigde technologieën 4, 5, 6, 7, 8, 9, chemiluminescente reporter dit systeem gebruik van een beeldvormende sonde die niet-radioactiev...

In de afgelopen decennia hebben beeldtechnologieën van de manier waarop artsen diagnosticeren en monitoren van de ziekte van een revolutie. Deze beeldvormende technieken zijn echter grotendeels beperkt tot gehele lichaam systemen, zoals positron emissie tomografie (PET), enkelvoudige foton-emissie-computertomografie (SPECT), computertomografie (CT) en magnetische resonantie imaging (MRI) is. Bijzondere aandacht is besteed aan kanker, en technologische doorbraken imaging zijn sterk verbeterd de manier waarop deze ziekte wordt gediagnosticeerd en behandeld. Ondanks deze vooruitgang, er is een plek waar deze beeldvormende technieken gewoon niet passen: de operatiekamer. Terwijl het gehele lichaam technieken helpen bij chirurgische planning, ze meestal onvoldoende ruimtelijke resolutie hoog genoeg om artsen te helpen bepalen realtime of alle tumorweefsel is verwijderd of resttumorweefsel verborgen blijft aan de snijvlakken 1. Ervoor zorgen dat er geen infiltratievetumor marges zijn achtergelaten is een van de belangrijkste chirurgische doelen, en chirurgen moeten een strakke-koord tussen strenge en voorzichtige weefsel resectie te lopen. Als er te veel wordt verwijderd, worden ongewenste bijwerkingen voor de patiënt vergroot; als er te weinig wordt verwijderd, worden recidief steeg met 2, 3. Daarom is het cruciaal om nauwkeurige tumorgrenzen bakenen, en wij geloven dat chemiluminescente intra-operatieve beeldvorming kan bijdragen tot de nauwkeurigheid van de identificatie van tumor marges te verbeteren door te helpen chirurgen kwaadaardig weefsel dat anders onopgemerkt bestaande technieken kunnen blijven visualiseren. Er zijn vele imaging technologieën momenteel onderzocht voor hun mogelijke bruikbaarheid als intra-operatieve imaging systemen. Deze omvatten β- en γ-straling-emitterende probes 4, 5 optische fluorescentie, Raman spectroscopie 6 </sup&gt;, 7 en Cherenkov luminescentie 8, 9. Tot dusver is geen van deze zijn vestigen standaard klinische instrumenten. Optische fluorescentiebeeldvorming dusver bewezen als de meest veelbelovende van deze technieken en is dus het meest onderzocht. Hoewel reeds aangetoond dat het een waardevol instrument voor vele toepassingen, is niet zonder beperkingen. Sterker nog, zijn belangrijkste nadeel is de achtergrond fluorescentie gegenereerd door inherent Autofluorescente biologisch weefsel. Deze achtergrond autofluorescente signaal is een product van de excitatie van het omringende weefsel, naast de fluorofoor door de externe lichtbron nodig voor het genereren van een fluorescerend signaal. Vanuit praktisch oogpunt kan deze autofluorescentie mogelijk leiden tot lage signaal-ruisverhoudingen, die het nut van deze technologie in de operatiekamer kan beperken. de voornaamstevoordeel van chemiluminescentie imaging via fluorescentiebeeldvorming is dat er geen excitatie licht nodig is. Daardoor is er geen achtergrond autofluorescentie. In chemiluminescentie beeldvorming wordt de excitatie-energie plaats chemisch gegenereerd. Deze werkwijze geeft geen ongewenste achtergrondsignaal en kan daarom leiden tot hogere signaal-ruisverhoudingen. Dit kan uiteindelijk leiden tot nauwkeuriger en nauwkeurige detectie van snijvlakken. Enigszins verrassend is het nut van deze aanpak als een intra-operatieve beeldvormende techniek onontgonnen 10 gebleven. Inderdaad, het dichtst voorbeeld van deze techniek is de oxidatie van luminol door myeloperoxidase in muizen 11, 12, 13. Chemiluminescente biomedische beeldvorming is daarmee tamelijk onontgonnen onderzoeksgebied dat de volgende voordelen kunnen bieden: (1) minimale autofluorescentie resulteert in een lage achtergrondsignaal met higher signal-to-noise ratio&#39;s; (2) afstembare golflengten van chemiluminescente emissie variëren van het zichtbare naar het nabije-infrarood; en (3) functionalizable chemoluminescente complexen die in combinatie met linker technologieën en gerichte biomoleculen reeds bestaande, toegang tot volledige bibliotheken van gerichte moleculaire beeldvorming sondes 14. Deze proof-of-principle studie toont het potentieel nut van chemiluminescent beeldvorming in de biomedische omgeving met behulp van een ruthenium-based imaging middel. De chemiluminescente eigenschappen van deze verbinding zijn goed bestudeerd, met onderzoeken die teruggaat tot het midden van de jaren 1960 15. Bij chemische activatie agens produceert het licht bij ongeveer 600 nm 16, die goed geschikt is voor medische beeldvorming doeleinden. De activeringsenergie wordt geleverd door een redoxreactie die leidt tot een aangeslagen toestand-die een levensduur van 650 ns in water 17 heeft -follverschuldigd door het genereren van fotonen op relaxatie van deze aangeslagen toestand. Door het gebruik van een speciaal ontworpen remote vernevelaar, waren we in staat om de verbinding zowel ex vivo en in vivo te detecteren. De resultaten van initiële experimenten zijn veelbelovend, suggereert verder onderzoek van deze technologie.

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			<td style="width:121px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:215px;">stainless steel rod (10 cm of 1/16&#8221; steel)</td>
			<td style="width:119px;">Metals Depot Int. Inc.</td>
			<td style="width:87px;">2192</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:215px;">Pencil Classic HB</td>
			<td style="width:119px;">Papermate</td>
			<td style="width:87px;">58592</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:215px;">Paper clip</td>
			<td style="width:119px;">Office Depot</td>
			<td style="width:87px;">221720</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:215px;">speaker cable</td>
			<td style="width:119px;">RCA Inc.</td>
			<td style="width:87px;">AH1650SN</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="39">
			<td height="39" style="height:39px;width:215px;">Energizer 9V alkaline battery</td>
			<td style="width:119px;">Energizer Holdings Inc.</td>
			<td style="width:87px;">EN22</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="61">
			<td height="61" style="height:61px;width:215px;">Hitech HS-82MG Micro Servo Motor, 3.4kg/cm output torque @ 6V</td>
			<td style="width:119px;">Hitech RCD USA Inc.</td>
			<td style="width:87px;">32082S</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:215px;">Name</td>
			<td style="width:119px;">Company</td>
			<td style="width:87px;">Catalog Number</td>
			<td style="width:121px;">Comments</td>
		</tr>
		<tr height="39">
			<td height="39" style="height:39px;width:215px;">28 cm plastic cable ties</td>
			<td style="width:119px;">General Electric Inc.</td>
			<td style="width:87px;">50725</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:215px;">Duct tape</td>
			<td style="width:119px;">3M Inc.</td>
			<td style="width:87px;">3939</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:215px;">littleBits w1 wire</td>
			<td style="width:119px;">littleBits Inc.</td>
			<td style="width:87px;">w1 wire</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:215px;">littleBits p1 power</td>
			<td style="width:119px;">littleBits Inc.</td>
			<td style="width:87px;">p1 power</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="39">
			<td height="39" style="height:39px;width:215px;">littleBits i2 toggle switch</td>
			<td style="width:119px;">littleBits Inc.</td>
			<td style="width:87px;">i2 toggle switch</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:215px;">littleBits 011 servo</td>
			<td style="width:119px;">littleBits Inc.</td>
			<td style="width:87px;">011 servo</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="41">
			<td height="41" style="height:41px;width:215px;">20 cm plastic covered wire twist ties</td>
			<td style="width:119px;">Four Star Plastics</td>
			<td style="width:87px;">71TIE8000</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="112">
			<td height="133" rowspan="2" style="height:133px;width:215px;">Tris(2,2&#8242;-bipyridyl)dichlororuthenium(II) hexahydrate</td>
			<td rowspan="2" style="width:119px;">Sigma-Aldrich Inc.</td>
			<td rowspan="2" style="width:87px;">224758</td>
			<td rowspan="2" style="width:121px;"></td>
		</tr>
		<tr height="21">
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:215px;">Ammonium cerium(IV) nitrate</td>
			<td style="width:119px;">Sigma-Aldrich Inc.</td>
			<td style="width:87px;">22249</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="39">
			<td height="39" style="height:39px;width:215px;">Isofluorane</td>
			<td style="width:119px;">Baxter Healthcare</td>
			<td style="width:87px;">1001936060</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:215px;">PBS</td>
			<td style="width:119px;">Sigma-Aldrich</td>
			<td style="width:87px;">PBS1</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:215px;">Ethanol</td>
			<td style="width:119px;">Sigma-Aldrich</td>
			<td style="width:87px;">2854</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:215px;">Triethylamine</td>
			<td style="width:119px;">Sigma-Aldrich Inc.</td>
			<td style="width:87px;">T0886</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="97">
			<td height="97" style="height:97px;width:215px;">Water</td>
			<td style="width:119px;"></td>
			<td style="width:87px;"></td>
			<td style="width:121px;">Water was purified using a Milipore Mili-Q (R &#8805; 18 M&#937;)</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:215px;">Female nude (outbred) mice</td>
			<td rowspan="2" style="width:119px;">Jackson Laboratories</td>
			<td rowspan="2" style="width:87px;">1929</td>
			<td rowspan="2" style="width:121px;">age 5 - 6 weeks</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:215px;">Strain C57BL/6J&#160;&#160;</td>
		</tr>
		<tr height="39">
			<td height="39" style="height:39px;width:215px;">NU/J male mice at&#160;</td>
			<td style="width:119px;">Jackson Laboratories</td>
			<td style="width:87px;">2019</td>
			<td style="width:121px;">age 6 &#8211; 8 weeks</td>
		</tr>
		<tr height="41">
			<td height="41" style="height:41px;width:215px;">IVIS 200 bioluminescence reader</td>
			<td style="width:119px;">Caliper Live Science</td>
			<td style="width:87px;"></td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:215px;">Live Image 4.2 software</td>
			<td style="width:119px;">Perkin-Elmer</td>
			<td style="width:87px;">128165</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:215px;">Microscope slides</td>
			<td style="width:119px;">ThermoScientific</td>
			<td style="width:87px;">4951PLUS4</td>
			<td style="width:121px;"></td>
		</tr>
	</tbody>
</table>

a novel technique for generating and observing chemiluminescence in a biological setting

Intraoperative imaging techniques have the potential to make surgical interventions safer and more effective; for these reasons, such techniques are quickly moving into the operating room. Here, we present a new approach that utilizes a technique not yet explored for intraoperative imaging: chemiluminescent imaging. This method employs a ruthenium-based chemiluminescent reporter along with a custom-built nebulizing system to produce ex vivo or in vivo images with high signal-to-noise ratios. The ruthenium-based reporter produces light following exposure to an aqueous oxidizing solution and re-reduction within the surrounding tissue. This method has allowed us to detect reporter concentrations as low as 6.9 pmol/cm2. In this work, we present a visual guide to our proof-of-concept in vivo studies involving subdermal and intravenous injections in mice. The results suggest that this technology is a promising candidate for further preclinical research and might ultimately become a useful tool in the operating room.

De vernevelaar systeem in dienst Protocol 1 omschreven kan worden opgebouwd uit gemakkelijk verkrijgbare materialen tegen een lage kostprijs. Het is bedoeld om een inzet op afstand getriggerde sproeien van het reducerende / oxiderende stof in een bioluminescente reader (figuur 1). Ons ontwerp zorgt voor een veilig gebruik van de vernevelaar in de bioluminescentie lezer op 14 cm afstand van de lens. Geen beslaan of vervaging van de lens werd waargenomen tijdens de operati...

Watch this Scientific Journal Video about A Novel Technique for Generating and Observing Chemiluminescence in a Biological Setting at JoVE.com

A Novel Technique for Generating and Observing Chemiluminescence in a Biological Setting

This protocol describes a new intraoperative imaging technique that uses a ruthenium complex as a source of chemiluminescent light emission, thereby producing high signal-to-noise ratios during in vivo imaging. Intraoperative imaging is an expanding field that could revolutionize the way that surgical procedures are performed.

Een nieuwe techniek voor het genereren en Observing Chemiluminescence in een biologische instelling

Intraoperative imaging techniques have the potential to make surgical interventions safer and more effective; for these reasons, such techniques are ...

Research

JoVE Journal

Bioengineering

A Method for Ovarian Follicle Encapsulation and Culture in a Proteolytically Degradable 3 Dimensional System

Een methode voor ovariële follikel Encapsulation en cultuur in een proteolytisch Afbreekbare 3 dimensionale systeem

The ovarian follicle is the functional unit of the ovary that secretes sex hormones and supports oocyte maturation. In vitro  follicle techniques provide ...

Fluorescent Nanoparticles for the Measurement of Ion Concentration in Biological Systems

Fluorescerende Nanodeeltjes voor het meten van Ion Concentratie in Biological Systems

Tightly regulated ion homeostasis throughout the body is necessary for the prevention of such debilitating states as dehydration.1 In contrast, rapid ion ...

Creating Two-Dimensional Patterned Substrates for Protein and Cell Confinement

Het creëren van twee-dimensionale Patterned Substraten voor eiwit en Cell opsluiting

Microcontact printing provides a rapid, highly reproducible method for the creation of well-defined patterned substrates.1  While microcontact printing ...

Cellular Lipid Extraction for Targeted Stable Isotope Dilution Liquid Chromatography-Mass Spectrometry Analysis

Cellulaire vetextractie voor gerichte stabiele isotopen Verdunning vloeistofchromatografie-massaspectrometrie Analyse

The metabolism of fatty acids, such as arachidonic acid (AA) and linoleic acid (LA), results in the formation of oxidized bioactive lipids, including ...

Density Gradient Multilayered Polymerization (DGMP): A Novel Technique for Creating Multi-compartment, Customizable Scaffolds for Tissue Engineering

Density Gradient Multilayered Polymerization DGMP: A Novel Technique for Creating Multi-compartment, Customizable Scaffolds for Tissue Engineering

Dichtheidsgradiënt Meerlaagse polymerisatie (dGMP): een nieuwe techniek voor het maken van meerdere compartimenten, Aanpasbare Steigers for Tissue Engineering

Complex tissue culture matrices, in which types and concentrations of biological stimuli (e.g. growth factors, inhibitors, or small molecules) or matrix ...

Observing and Quantifying Fibroblast-mediated Fibrin Gel Compaction

Observeren en kwantificeren van fibroblast-gemedieerde Fibrin Gel Compaction

Cells embedded in collagen and fibrin gels attach and exert traction forces on the fibers of the gel. These forces can lead to local and global ...

A Novel Stretching Platform for Applications in Cell and Tissue Mechanobiology

A Novel Stretching platform voor toepassingen in de Cel en Weefsel Mechanobiology

Tools that allow the application of mechanical forces to cells and tissues or that can quantify the mechanical properties of biological tissues have ...

Bioprinting of Cartilage and Skin Tissue Analogs Utilizing a Novel Passive Mixing Unit Technique for Bioink Precellularization

Bioprinting van kraakbeen en huid weefsel analogen met behulp van een roman passief mengen eenheid techniek voor Bioink Precellularization

Bioprinting is a powerful technique for the rapid and reproducible fabrication of constructs for tissue engineering applications. In this study, both ...

Microdissection of Primary Renal Tissue Segments and Incorporation with Novel Scaffold-free Construct Technology

Microdissection van primaire renale weefsel segmenten en integratie met nieuwe steiger-vrije constructie technologie

Kidney transplantation is now a mainstream therapy for end-stage renal disease. However, with approximately 96,000 people on the waiting list and only ...