Name: Intravitale subcellulaire microscopie van de borstklier (Video) | JoVE Journal
Uploaded: 2022-06-24T14:00:00.000+00:00
Duration: 4 min 14 s
Description: Leer intravitale subcellulaire microscopie van de borstklier met laser scanning confocale en multiphoton microscopie. Deze methode maakt real-time visualisatie van secretorische epitheelcellen en lipidedruppels mogelijk.

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</ol>

<table><tbody><tr><td>488 laser</td><td>Melles-Griot</td><td>-</td><td>CW&nbsp; laser 50 mW</td></tr><tr><td>60x PLAPON oil immersion objective (NA 1.42)</td><td>Olympus</td><td>1-U2B933</td><td>Lens Confocal microscope</td></tr><tr><td>633 laser</td><td>Melles-Griot</td><td>-</td><td>CW He-Ne laser 12 mW</td></tr><tr><td>63x objective (NA 1.40, HC PL APO CS2)</td><td>Leica</td><td>11506350</td><td>Lens Two-photon microscope</td></tr><tr><td>BA 410-460 nm</td><td>Chroma</td><td>-</td><td>Band-pass filter</td></tr><tr><td>BA 495-540 nm</td><td>Chroma</td><td>-</td><td>Band-pass filter</td></tr><tr><td>BA 505-605 nm</td><td>Chroma</td><td>-</td><td>Band-pass filter</td></tr><tr><td>BA 655-755 nm</td><td>Chroma</td><td>-</td><td>Band-pass filter</td></tr><tr><td>Boron-dipyrromethane (BODIPY) 665/676</td><td>Thermo Fisher Scientific</td><td>B3932</td><td>Lipid peroxiation sensor</td></tr><tr><td>Carbomer-940</td><td>Snowdrift Farm</td><td>739601480651</td><td>Gel</td></tr><tr><td>Catheter</td><td>Terumo</td><td>SV27EL</td><td>Winged infusion sets&nbsp;</td></tr><tr><td>Cauterizer&nbsp;</td><td>Braintree Scientific, Inc</td><td>GEM 5917</td><td>Cautery system</td></tr><tr><td>CMV-Cre mouse&nbsp;</td><td>Jackson lab</td><td>006054</td><td>Mouse line</td></tr><tr><td>Coverslip</td><td>Bioptechs</td><td>-</td><td>30mm diameter coverlip for inverted microscope</td></tr><tr><td>Curity 4x4 inch all purpose sponge gauze</td><td>Covidien</td><td>9024</td><td>Sponge</td></tr><tr><td>EGFPcyto mouse</td><td>Jackson lab</td><td>003291</td><td>Mouse line</td></tr><tr><td>Fiji/ImageJ software</td><td>Open source</td><td>-</td><td>Free software tool</td></tr><tr><td>Fine forceps</td><td>Braintree Scientific, Inc</td><td>FC003 8</td><td>Tissue forceps</td></tr><tr><td>Fluoview 1000 microscope</td><td>Olympus</td><td>FV1000</td><td>Confocal microscope</td></tr><tr><td>FluoView software</td><td>Olympus</td><td>-</td><td>Confocal microscope and Two-photon microscope</td></tr><tr><td>Hand-held electric razor</td><td>Braintree Scientific, Inc</td><td>CLP-8786-451A</td><td>Cordless clipper</td></tr><tr><td>Heat pad</td><td>Braintree Scientific, Inc</td><td>DPIP</td><td>Heat pad for animals</td></tr><tr><td>HyD detectors</td><td>Leica</td><td>-</td><td>Leica 4Tune spectral detector</td></tr><tr><td>Imaris software</td><td>Bitplane / Oxford instruments</td><td>-</td><td>Commercial software tool</td></tr><tr><td>Ingisht X3 tunable laser</td><td>Spectra Physics</td><td>Insight X3</td><td>Tunable Pulse-Laser</td></tr><tr><td>Isoflurane</td><td>VetOne</td><td>13985-046-40</td><td>Anesthetic</td></tr><tr><td>Ketamine&nbsp;</td><td>VetOne</td><td>13985-702-10</td><td>Anesthetic</td></tr><tr><td>LAS X Software</td><td>Leica</td><td>-</td><td>Two-photon microscope software tool</td></tr><tr><td>Mai-Tai tunable laser</td><td>Spectra Physics</td><td>Mai-Tai</td><td>Laser</td></tr><tr><td>MetaMorph</td><td>Molecular Devices</td><td>-</td><td>Commercial software tool</td></tr><tr><td>Monodansylpentane AUTODOT</td><td>Abcepta</td><td>Sm1000a</td><td>Lipid droplet dye</td></tr><tr><td>MPE-RS microscope</td><td>Olympus</td><td>IX70</td><td>Two-photon microscope</td></tr><tr><td>mT/mG mouse</td><td>Jackson lab</td><td>007676</td><td>Mouse line</td></tr><tr><td>Objective heater</td><td>Bioptechs</td><td>150819</td><td>Objective heater for both confocal and two-photon microscopes</td></tr><tr><td>Oxygen-saturated respiration chamber</td><td>Patterson Scientific</td><td>78933385, SAS3 and EVAC4</td><td>Gas Anesthesia and evacuation system&nbsp;</td></tr><tr><td>Parafilm</td><td>Heathrow Scientific</td><td>HS234526B</td><td>Semi-transparent, flexible, thermoplastic film</td></tr><tr><td>PMT detector</td><td>Olympus</td><td>-</td><td>Descanned&nbsp;&nbsp; detectors</td></tr><tr><td>PMT detector</td><td>LSM-Technology</td><td>Custom built</td><td>Non-Descanned Detectors</td></tr><tr><td>Pump</td><td>Harvard Apparatus</td><td>703602, 704402</td><td>Nanomite injector and controller</td></tr><tr><td>Saline</td><td>Quality Biological</td><td>114-055-721EA</td><td>Normal saline</td></tr><tr><td>Sharp blunt-ended scissors</td><td>Braintree Scientific, Inc</td><td>SCT-S 508</td><td>Surgical scissors</td></tr><tr><td>Syringe</td><td>Covidien</td><td>22-257-150</td><td>1mL tuberculin syringe</td></tr><tr><td>TCS SP8 Dive Spectral microscope</td><td>Leica</td><td>SP8</td><td>Two-photon microscope</td></tr><tr><td>Tweezers&nbsp;</td><td>Braintree Scientific, Inc</td><td>FC032</td><td>Tissue forceps</td></tr><tr><td>Xylazine&nbsp;</td><td>VetOne</td><td>13985-704-10</td><td>Anesthetic</td></tr></tbody></table>

intravitale subcellulaire microscopie van de borstklier

Intravital Subcellular Microscopy of the Mammary Gland

Intravitale subcellulaire microscopie van de borstklier

The mammary gland constitutes a model par excellence for investigating epithelial functions, including tissue remodeling, cell polarity, and secretory ...

intravital-subcellular-microscopy-of-the-mammary-gland

Research

JoVE Journal

Biology

Biologie

Time-lapse imaging wordt gebruikt om het gedrag van primaire preneoplastische mammaire epitheelcellen afkomstig van genetisch gemanipuleerde muizen met borstkanker te bepalen of er correlaties tussen specifieke gedragsparameters en verschillende genetische laesies beoordelen.

Time-lapse imaging of Primary preneoplastische uier epitheelcellen afkomstig van de genetisch gemanipuleerde muis modellen van borstkanker

Opklaren is een krachtige tool waarmee beeldvorming verschillende biologische processen in levende dieren. In dit artikel presenteren we een gedetailleerde werkwijze voor het afbeelden van de dynamiek van subcellulaire structuren, zoals de secretoire granules in de speekselklieren van levende muizen.

Intravitale Microscopy for Imaging Subcellulaire Structuren in Levende muizen die fluorescerende eiwitten

Een protocol voor de niet-invasieve intraductale afgifte van waterige reagentia voor de muis borstklier wordt beschreven. De methode maakt gebruik van gelokaliseerde injectie in de tepels van melkklieren targeting mamma-kanalen specifiek. Deze techniek kan worden aangepast voor verschillende verbindingen waaronder siRNA, chemotherapeutische middelen en kleine moleculen.

Intraductale Injectie voor Gelokaliseerde Drug Delivery naar de muis borstklier

De indirecte immunofluorescentie protocol beschreven in dit artikel laat de detectie en de lokalisatie van eiwitten in de muis borstklier. Een complete methode wordt gegeven voor te bereiden borstklier monsters, om immunohistochemie uit te voeren, om de afbeelding van het weefsel secties met behulp van fluorescentie microscopie, en om beelden te reconstrueren.

Indirecte Immunofluorescentie op Bevroren Secties van Muis borstklier

Dit protocol beschrijft het gebruik van multiphoton microscopie tot uitgebreide time-lapse beeldvorming van meercellige interacties uit te voeren in real time, in vivo bij enkele resolutie cel.

Extended Time-lapse intravitale Imaging van Real-time Meercellige Dynamiek in de tumor micro

Hier beschrijven we een techniek voor de gelokaliseerde toediening van reagentia aan de borstklier van konijnen via een intraductale injectie. Daarnaast beschrijven we een protocol voor visualisatie en bevestiging van de toediening door middel van hoogwaardige echografie met contrastmiddelen.

Intraductale Levering aan de Rabbit borstklier

Hier presenteren we een protocol voor de isolatie van geheel, intact muis borstklieren om te onderzoeken van de extracellulaire matrix (ECM) expressie en ductaal morfologie. Muis #4 abdominale klieren werden gehaald uit de 8-10 week oude vrouwelijke nullipaar muizen, vast in neutraal gebufferde formaline, gesegmenteerd en gekleurd met behulp van immunohistochemistry voor ECM eiwitten.

Isolatie van Intact, hele muis borstklieren p.a. van extracellulaire Matrix expressie en klier morfologie

Hier wordt een benadering gepresenteerd voor intravitale beeldvorming op lange termijn met behulp van optisch heldere, siliconen vensters die rechtstreeks op het weefsel / orgaan van belang en de huid kunnen worden gelijmd. Deze ramen zijn goedkoper en veelzijdiger dan andere die momenteel in het veld worden gebruikt, en de chirurgische inbrenging veroorzaakt beperkte ontsteking en leed voor de dieren.

Longitudinale intravitale beeldvorming door heldere siliconenvensters

Dit protocol beschrijft een nieuw mammabeeldvormingsvenster met een vervangbaar deksel (R.MIW). Intravitale microscopie na implantatie van de R.MIW maakt longitudinale en meerdaagse beeldvorming van de gezonde en zieke borstklier mogelijk met een cellulaire resolutie tijdens de verschillende ontwikkelingsstadia.

Longitudinale intravitale microscopie met behulp van een mammabeeldvormingsvenster met vervangbaar deksel

De hier beschreven intravitale beeldvormingsmethode maakt gebruik van collageen tweede harmonische generatie en endogene fluorescentie van de metabole co-factor NAD (P) H om een niet-invasieve tumormicro-omgeving te segmenteren in tumor-, stromale en vasculaire compartimenten voor diepgaande analyse van 4D intravitale beelden.

Een labelvrije segmentatiebenadering voor intravitale beeldvorming van de micro-omgeving van borsttumoren

Het huidige protocol beschrijft intraductale injectie van virale vectoren via de speen om genen van belang af te leveren in de borstepitheelcellen.

In Vivo Genafgifte in muisepitheelcellen via intraductale injectie van zmammas

Leer intravitale subcellulaire microscopie van de borstklier met laser scanning confocale en multiphoton microscopie. Deze methode maakt real-time visualisatie van secretorische epitheelcellen en lipidedruppels mogelijk.

Intravitale subcellulaire microscopie van de borstklier

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