Name: catalytic scavenging of plant reactive oxygen species in vivo by anionic cerium oxide nanoparticles
Uploaded: 2018-08-26T15:00:00
Description: Watch this Scientific Journal Video about Catalytic Scavenging of Plant Reactive Oxygen Species In Vivo by Anionic Cerium Oxide Nanoparticles at JoVE.com

Dette arbeidet ble støttet av University of California, Riverside og USDA National Institute of Food og landbruk, Luke prosjektet 1009710 J.P.G. Dette materialet er basert på arbeid støttes av National Science Foundation under Grant nr. 1817363 til J.P.G.

1. voksende A. thaliana planter
<ol><li>Så A. thaliana frø i 5 cm x 5 cm disponibel potter fylt med standard jord blanding. Sette 32 av disse potter i en plast brett fylt med vann (~ 0,5 cm dybde) og overføre det plast brettet med planter i en plante vekst kammer.<ol>
<li>Angi veksten kammer innstillingene som følger: 200 µmol/ms fotosynteseaktiviteten aktiv stråling (PAR), 24 ± 1 ° C dag og 21 ± 1 ° C natt, 60% fukt og 14/10 h dag/natt lyset regimet, henholdsvis.</li>
	</ol>
</li>
	<li>Tynn...

<ol>
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I denne protokollen beskriver vi PNC syntese, karakterisering, fluorescerende farge merking og AC confocal avbildning av nanopartikler i anlegget mesophyll celler viser i vivo ROS scavenging aktiviteten deres. PNC er synthesized fra en blanding av cerium nitrat og PAA løsning i ammonium hydroksid. PNC kjennetegnes av absorpsjon spectrophotomery og konsentrasjonen fastsatt ved bruk av øl-Lamberts lov. Zeta potensielle mål bekreftet negativt ladde overflaten av PNC for å forbedre levering til chloroplasts<sup ...

Cerium-oksid nanopartikler (nanoceria) er mye brukt i levende organismer, fra grunnleggende forskning til bioteknologi, på grunn av deres forskjellige katalytisk reaktive oksygen arter (ROS) scavenging evne til1,2,3. Nanoceria har ROS scavenging evner på grunn av et stort antall overflaten oksygen stillinger som veksler mellom to oksidasjon stater (Ce3 + og Ce4 +) 4,5,6. Ce3 + dingler obligasjoner renovere effektivt ROS mens gitter påkjenninger på nanoskala fremme regenereringen av disse defekt nettsteder via redoks sykling reaksjoner7. Nanoceria har også blitt nylig brukt for å studere og engineering plante funksjon8,9. Planter under abiotiske stress oppleve opphopning av ROS, forårsaker oksidative skader lipider, proteiner og DNA10. I A. thaliana planter fører nanoceria katalytisk renovering av ROS i vivo til bedre fotosyntese under høy lys, varme og kjøling påkjenninger8. Bruk nanoceria jord også øker skyte biomasse og korn avkastning av hvete (Triticum aestivum)11; raps (Brassica napus) planter behandlet med nanoceria har høyere anlegget biomasse under salt stress12.
Nanoceria tilbyr bioingeniører og plante biologer en nanoteknologi-basert verktøy for å forstå abiotiske stressresponser og forbedre anlegget abiotiske stress toleranse. Nanocerias i vivo ROS scavenging evner er uavhengig av plantearter og lettvinte levering i anlegget vev har potensial til å aktivere bred anvendelse utenfor modell organismer. I motsetning til andre genetisk-baserte metoder krever ikke nanoceria generere anlegget linjer med overuttrykte av antioksidant enzymer for høyere ROS scavenging evne til13. Blad lamina infiltrasjon av nanoceria planter er en praktisk tilnærming til lab-basert forskning.
Det overordnede målet med denne protokollen er å beskrive 1) syntese og karakterisering av negativt ladde poly (akryl) syre nanoceria (PNC), 2) levering og sporing av PNC hele bladet cellene, og 3) overvåking av PNC-aktiverte ROS scavenging i vivo. I denne protokollen, negativt ladde poly (akryl) syre nanoceria (PNC) syntetisert og preget av deres absorpsjon spektrum, etter diameter og zeta potensielle. Vi beskriver en enkelt blad lamina infiltrasjon metode for å levere PNC i anlegget blad vev. I vivo avbildning av hydrogenion distribusjon i mesophyll celler, var en fluorescerende farge (DiI) pleide å merke PNC (DiI-PNC) og observere nanopartikler via AC confocal fluorescens mikroskopi. Til slutt forklare vi hvordan overvåke i vivo PNC ROS scavenging gjennom AC confocal mikroskopi.

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			<td height="21" style="height:21px;width:567px;">Cerium (III) nitrate hexahydrate</td>
			<td style="width:228px;">Sigma-Aldrich</td>
			<td style="width:165px;">238538-100G</td>
			<td style="width:265px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">Molecular Biology Grade Water, Corning</td>
			<td style="width:228px;">VWR</td>
			<td style="width:165px;">45001-044&#160;</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">Falcon 50 mL Conical Centrifuge Tubes</td>
			<td style="width:228px;">VWR</td>
			<td style="width:165px;">14-959-49A</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">Poly (acrylic acid) 1,800 Mw</td>
			<td style="width:228px;">Sigma-Aldrich</td>
			<td style="width:165px;">323667-100G</td>
			<td></td>
		</tr>
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			<td height="21" style="height:21px;width:567px;">Fisherbrand Digital Vortex Mixer</td>
			<td style="width:228px;">Fisher Scientific</td>
			<td style="width:165px;">02-215-370</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">Fisherbrand Digital Vortex Mixer Accessory, Insert Retainer</td>
			<td style="width:228px;">Fisher Scientific</td>
			<td style="width:165px;">02-215-391</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">Fisherbrand Digital Vortex Mixer Accessories: Foam Insert Set</td>
			<td style="width:228px;">Fisher Scientific</td>
			<td style="width:165px;">02-215-395</td>
			<td></td>
		</tr>
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			<td height="21" style="height:21px;width:567px;">Ammonium hydroxide solution</td>
			<td style="width:228px;">Sigma-Aldrich</td>
			<td style="width:165px;">05002-1L</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">PYREX Griffin Beakers, Graduated, Corning</td>
			<td style="width:228px;">VWR</td>
			<td style="width:165px;">13912-149&#160;</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:567px;">RCT basic</td>
			<td style="width:228px;">IKA</td>
			<td style="width:165px;">3810001</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">Eppendorf Microcentrifuge 5424</td>
			<td style="width:228px;">VWR</td>
			<td style="width:165px;">80094-126</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">Amicon Ultra-15 Centrifugal Filter Units</td>
			<td style="width:228px;">Millipore-Sigma</td>
			<td style="width:165px;">UFC901024</td>
			<td></td>
		</tr>
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			<td height="21" style="height:21px;width:567px;">Allegra X-30 Series Benchtop Centrifuge</td>
			<td style="width:228px;">Beckman Coulter</td>
			<td style="width:165px;">B06314</td>
			<td></td>
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			<td height="21" style="height:21px;width:567px;">UV-2600 Sptecrophotometer</td>
			<td style="width:228px;">Shimadzu</td>
			<td style="width:165px;">UV-2600 120V</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:567px;">Whatman Anotop 10 syringe filter</td>
			<td style="width:228px;">Sigma-Aldrich</td>
			<td style="width:165px;">WHA68091102</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">BD Disposable Syringes with Luer-Lok Tips</td>
			<td style="width:228px;">Fisher Scientific</td>
			<td style="width:165px;">14-829-45</td>
			<td></td>
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			<td height="21" style="height:21px;width:567px;">Zetasizer Nano S</td>
			<td style="width:228px;">Malvern Panalytical</td>
			<td style="width:165px;">Zen 1600</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">1,1&#8242;-Dioctadecyl-3,3,3&#8242;,3&#8242;-tetramethylindocarbocyanine perchlorate</td>
			<td style="width:228px;">Sigma-Aldrich</td>
			<td style="width:165px;">42364-100MG</td>
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			<td height="21" style="height:21px;width:567px;">Dimethyl Sulfoxide, ACS</td>
			<td style="width:228px;">VWR</td>
			<td style="width:165px;">BDH1115-1LP</td>
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			<td height="21" style="height:21px;width:567px;">Sunshine Mix #1 LC1</td>
			<td style="width:228px;">Green Island Distributors, Inc</td>
			<td style="width:165px;">5212601.CFL080P</td>
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			<td height="21" style="height:21px;width:567px;">Adaptis 1000</td>
			<td style="width:228px;">Conviron</td>
			<td style="width:165px;">A1000</td>
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			<td height="21" style="height:21px;width:567px;">TES, &#62;99% (titration</td>
			<td style="width:228px;">Sigma-Aldrich</td>
			<td style="width:165px;">T1375-100G</td>
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			<td height="21" style="height:21px;width:567px;">Magnesium chloride</td>
			<td style="width:228px;">Sigma-Aldrich</td>
			<td style="width:165px;">M8266-1KG</td>
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			<td height="21" style="height:21px;width:567px;">Air-Tite All-Plastic Norm-Ject Syringe</td>
			<td style="width:228px;">Fisher Scientific</td>
			<td style="width:165px;">14-817-25</td>
			<td></td>
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			<td height="21" style="height:21px;width:567px;">Kimberly-Clark Professional Kimtech Science Kimwipes Delicate Task Wipers</td>
			<td style="width:228px;">Fisher Scientific</td>
			<td style="width:165px;">06-666A</td>
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			<td height="21" style="height:21px;width:567px;">Carolina Observation Gel</td>
			<td style="width:228px;">Carolina</td>
			<td style="width:165px;">132700</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">Corning microscope slides, frosted one side, one end</td>
			<td style="width:228px;">Sigma-Aldrich</td>
			<td style="width:165px;">CLS294875X25-72EA</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">Cork Borer Sets with Handles</td>
			<td style="width:228px;">Fisher Scientific</td>
			<td style="width:165px;">S50166A</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">Perfluorodecalin</td>
			<td style="width:228px;">Sigma-Aldrich</td>
			<td style="width:165px;">P9900-25G</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">Micro Cover Glasses, Square, No. 1</td>
			<td style="width:228px;">VWR</td>
			<td style="width:165px;">48366-045</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">Leica Laser Scanning Confocal Microscope TCS SP5</td>
			<td style="width:228px;">Leica Microsystems</td>
			<td style="width:165px;">TCS SP5</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">2&#8242;,7&#8242;-Dichlorofluorescin diacetate</td>
			<td style="width:228px;">Sigma-Aldrich</td>
			<td style="width:165px;">D6883-250MG</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">Dihydroethidium</td>
			<td style="width:228px;">Sigma-Aldrich</td>
			<td style="width:165px;">D7008-10MG</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">Fisherbrand Premium Microcentrifuge Tubes: 1.5 mL</td>
			<td style="width:228px;">Fisher Scientific</td>
			<td style="width:165px;">05-408-129</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">Eppendorf Uvette cuvettes</td>
			<td style="width:228px;">Sigma-Aldrich</td>
			<td style="width:165px;">Z605050-80EA</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:567px;">Chlorophyll meter&#160;</td>
			<td style="width:228px;">Konica Minolta</td>
			<td style="width:165px;">SPAD-502</td>
			<td></td>
		</tr>
	</tbody>
</table>

catalytic scavenging of plant reactive oxygen species in vivo by anionic cerium oxide nanoparticles

Reaktive oksygen arter (ROS) akkumulering er et kjennetegn på anlegget abiotiske stressrespons. ROS spille en dobbel rolle i planter ved å opptre som signalnettverk molekyler på lavt nivå og ødeleggende molekyler på høye nivåer. Akkumulering av ROS i stresset planter kan skade metabolitter, enzymer, lipider og DNA, forårsaker en reduksjon av plantevekst og avkastning. Evne til cerium oksid nanopartikler (nanoceria) katalytisk åtseleter ROS i vivo tilbyr unikt verktøy å forstå og bioengineer anlegget abiotiske stress toleranse. Her presenterer vi en protokoll for å syntetisere og karakterisere poly (akryl) syre belagt nanoceria (PNC), grensesnitt nanopartikler med planter via blad lamina infiltrasjon og overvåke deres distribusjon og ROS scavenging i vivo med AC confocal mikroskopi. Gjeldende molekylære verktøy for å manipulere ROS akkumulering i planter er begrenset til modell arter og krever arbeidskrevende transformasjon metoder. Denne protokollen for i vivo ROS scavenging har potensial til å bli brukt på wild type planter med store blader og blad struktur som Arabidopsis thaliana.

PNC syntese og karakterisering . PNC ble syntetisert, renset og preget følgende metoden beskrevet i protokollen del 2. Figur 1 En viser fargeleggingen løsninger av cerium nitrat, PAA, blandingen av cerium nitrat og PAA og PNC. En fargeendring fra hvit til lys gul vises etter PNC er syntetisert. Etter rensing med en 10 kDa filter, PNC var preget med et UV-VIS spektrofotometer. En toppen av absorb...

Watch this Scientific Journal Video about Catalytic Scavenging of Plant Reactive Oxygen Species In Vivo by Anionic Cerium Oxide Nanoparticles at JoVE.com

Catalytic Scavenging of Plant Reactive Oxygen Species In Vivo by Anionic Cerium Oxide Nanoparticles

Katalytisk renovering av anlegget reaktive oksygen arter i Vivo av Anionic Cerium oksid nanopartikler

Her presenterer vi en protokoll for syntese og karakterisering av cerium oksid nanopartikler (nanoceria) for ROS (reaktive oksygen arter) scavenging i vivo, nanoceria imaging i anlegget vev av AC confocal mikroskopi og i vivo overvåking av nanoceria ROS scavenging av AC confocal mikroskopi.

Reactive oxygen species (ROS) accumulation is a hallmark of plant abiotic stress response. ROS play&#160;a dual role in plants by acting as signaling ...

Research

JoVE Journal

Bioengineering

Catalytic Scavenging of Plant Reactive Oxygen Species In Vivo by Anionic Cerium Oxide Nanoparticles

A Protocol for Detecting and Scavenging Gas-phase Free Radicals in Mainstream Cigarette Smoke

En protokoll for å oppdage og scavenging Gass-fase frie radikaler i Mainstream sigarettrøyk

Cigarette smoking is associated with human cancers. It has been reported that most of the lung cancer deaths are caused by cigarette smoking 5,6,7,12.  ...

Viral Nanoparticles for In vivo Tumor Imaging

Viral Nanoparticles for In vivo Tumor Imaging

Viral Nanopartikler for<em&gt; In vivo</em&gt; Tumor Imaging

The use of nanomaterials has the potential to revolutionize materials science and medicine. Currently, a number of different nanoparticles are being ...

Fabrication of a Functionalized Magnetic Bacterial Nanocellulose with Iron Oxide Nanoparticles

Fabrikasjon av en funksjon Magnetic Bakteriell Nanocellulose med jernoksid nanopartikler

In this study, bacterial nanocellulose (BNC) produced by the bacteria Gluconacetobacter xylinus is synthesized and impregnated in situ with iron oxide ...

Cell Labeling and Targeting with Superparamagnetic Iron Oxide Nanoparticles

Cell Merking og målretting med superparamagnetiske Iron Oxide Nanopartikler

Targeted delivery of cells and therapeutic agents would benefit a wide range of biomedical applications by concentrating the therapeutic effect at the ...

Identification of Metal Oxide Nanoparticles in Histological Samples by Enhanced Darkfield Microscopy and Hyperspectral Mapping

Identifisering av Metal Oxide Nanopartikler i Histologiske Samples av Enhanced Darkfield Mikroskopi og Hyperspektral Mapping

Nanomaterials are increasingly prevalent throughout industry, manufacturing, and biomedical research. The need for tools and techniques that aid in the ...

Microwave-driven Synthesis of Iron Oxide Nanoparticles for Fast Detection of Atherosclerosis

Mikrobølgeovn-drevet Syntese av Iron Oxide Nanopartikler for Fast Påvisning av åreforkalkning

A fast and reproducible microwave-driven protocol has been developed for the synthesis of neridronate-functionalized nanoparticles. Starting from the ...

Production of Metal Nanoparticles by Pulsed Laser-ablation in Liquids: A Tool for Studying the Antibacterial Properties of Nanoparticles

Produksjon av metall nanopartikler ved pulserende laserablation i væsker: Et verktøy for å studere de antibakterielle egenskapene til nanopartikler

The emergence of multidrug-resistant bacteria is a global clinical concern leading some to speculate about our return to a "pre-antibiotics" era of ...

Rigid Embedding of Fixed and Stained, Whole, Millimeter-Scale Specimens for Section-free 3D Histology by Micro-Computed Tomography

Stive innebygging av fast og farget, hele, Millimeter skala prøver for delen-gratis 3D Histology av mikro-beregnet tomografi

For over a hundred years, the histological study of tissues has been the gold standard for medical diagnosis because histology allows all cell types in ...

Detection of Endotoxin in Nano-formulations Using Limulus Amoebocyte Lysate (LAL) Assays

Detection of Endotoxin in Nano-formulations Using Limulus Amoebocyte Lysate LAL Assays

Deteksjon av Endotoxin i Nano-formuleringer bruker Limulus Amoebocyte Lysate (LAL) analyser

When present in pharmaceutical products, a Gram-negative bacterial cell wall component endotoxin (often also called lipopolysaccharide) can cause ...

Using Magnetometry to Monitor Cellular Incorporation and Subsequent Biodegradation of Chemically Synthetized Iron Oxide Nanoparticles

Bruke magnetometry til å overvåke cellulær inkorporering og påfølgende biologisk nedbrytning av kjemisk synthetiserte jernoksid nanopartikler

Magnetic nanoparticles, made of iron oxide, present a peculiar interest for a wide range of biomedical applications for which they are often internalized ...