Name: Microscopia Subcelular Intravital da Glândula Mamária (Vídeo) | JoVE Journal
Uploaded: 2022-06-24T14:00:00.000+00:00
Duration: 4 min 14 s
Description: Aprenda microscopia confocal e multiphoton intravital da glândula mamária lactante em camundongos. Este método permite visualizar dinâmica de gotículas lipídicas e morfologia glandular em tempo real.

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</ol>

<table><tbody><tr><td>488 laser</td><td>Melles-Griot-CW</td><td></td><td>&nbsp; laser 50 mW</td></tr><tr><td>60x PLAPON objetiva de imersão em óleo (NA 1.42)</td><td>Olympus</td><td>1-U2B933</td><td>Lente Microscópio confocal</td></tr><tr><td>633 laser</td><td>Melles-Griot-CW</td><td></td> <td>He-Ne laser 12 mW</td></tr><tr><td>63x objetiva (NA 1.40, HC PL APO CS2)</td><td>Lente Leica</td><td>11506350</td><td>Microscópio de dois fótons</td></tr><tr><td>BA 410-460 nm</td><td>Chroma</td><td>-</td><td>Filtro passa-banda</td></tr><tr><td>BA 495-540 nm</td> <td>Filtro passa-banda</td><td>cromado</td><td></td></tr> <tr><td>BA 505-605 nm</td> <td>Filtro passa-banda</td><td>cromado</td><td></td></tr> <tr><td>BA 655-755 nm</td> <td>Filtro passa-banda</td><td></td> <td>cromado</td></tr> <tr><td>Boro-dipirrometano (BODIPY) 665/676</td><td>Thermo Fisher Scientific</td><td>B3932</td><td>Sensor de peroxiação lipídica</td></tr><tr><td>Carbomer-940</td><td>Snowdrift Farm</td><td>Cateter de</td><td>Gel</td>739601480651</tr><tr><td></td><td>Terumo</td><td>SV27EL</td><td>Conjuntos de infusão com asas&nbsp;</td></tr><tr><td>Cauterizador</td><td>Braintree Scientific, Inc</td><td>GEM 5917</td><td>Sistema de cauterização</td></tr><tr><td>CMV-Cre mouse&nbsp;</td><td>Jackson lab</td><td>006054</td><td>linha de camundongos</td></tr><tr><td>Coverslip</td><td>Bioptechs-30mm</td><td></td> <td>de diâmetro coverlip para microscópio invertido</td></tr><tr><td>Curity 4x4 polegadas gaze esponja multiuso</td><td>Covidien</td><td>9024</td><td>Esponja</td></tr><tr><td>EGFPcyto mouse</td><td>Jackson lab</td><td>003291</td><td>linha de mouse</td></tr><tr><td>Fiji / ImageJ software</td><td>de código</td> <td>aberto-ferramenta</td> <td>de software livre</td></tr><tr><td>Pinça fina</td><td>Braintree Scientific, Inc</td><td>FC003 8</td><td>Pinça de tecido</td></tr><tr><td>Microscópio Fluoview 1000 Microscópio</td><td>confocal</td><td>Olympus</td><td>FV1000</td></tr><tr><td>Software FluoView</td> <td>Microscópio Olympus-Confocal e microscópio de dois fótons</td><td></td><td></td></tr> <tr><td>Barbeador elétrico portátil</td><td>Braintree Scientific, Inc</td><td>CLP-8786-451A</td><td>Cortador sem fio</td></tr><tr><td>Almofada térmica</td><td>Braintree Scientific, Inc</td><td>DPIP</td><td>Almofada térmica para animais</td></tr><tr><td>Detectores HyD</td><td></td> <td>Leica-Leica</td> <td>4Tune detector espectral</td></tr><tr><td>Software Imaris Bitplane</td><td>/ Oxford instruments-Commercial</td><td></td> <td>ferramenta de software</td></tr><tr><td>Ingisht X3 laser sintonizável</td><td>Spectra Physics</td><td>Insight X3</td><td>Laser de pulso ajustável</td></tr><tr><td>Isoflurano</td><td>VetOne</td><td>13985-046-40</td><td>Anestésico</td></tr><tr><td>Cetamina&nbsp;</td><td>VetOne</td><td>13985-702-10</td><td>Anestésico</td></tr><tr><td>LAS X Software</td><td>Leica-Ferramenta</td> <td>de</td> <td>software de microscópio de dois fótons</td></tr><tr><td>Mai-Tai laser sintonizável Spectra</td><td>Physics</td><td>Mai-Tai</td><td>Laser</td></tr><tr><td>MetaMorph</td><td>Molecular Devices-Commercial</td><td></td> <td>ferramenta de software</td></tr><tr><td>Monodansilpentano AUTODOT</td><td>Abcepta</td><td>Sm1000a</td><td>Corante de gotículas lipídicas</td></tr><tr><td>MPE-RS microscópio</td><td>Olympus</td><td>IX70</td><td>Microscópio de dois fótons</td></tr><tr><td>mT/mG mouse</td><td>Jackson lab</td><td>007676</td><td>Linha de mouse</td></tr><tr><td>Aquecedor de objetiva</td><td>Bioptechs</td><td>150819</td><td>Aquecedor de objetiva para microscópios confocais e de dois fótons</td></tr><tr><td>Câmara de respiração saturada de oxigênio</td><td>Patterson Scientific</td><td>78933385, SAS3 e EVAC4</td><td>Sistema de anestesia e evacuação a gás&nbsp;</td></tr><tr><td>Parafilm</td><td>Heathrow Scientific</td><td>HS234526B</td><td>Detector PMT de filme termoplástico semi-transparente, flexível e transparente</td></tr><tr><td></td><td></td> Olympus-Descanned<td></td><td>&nbsp; &nbsp; detectores</td></tr><tr><td>detector PMT</td><td>LSM-Technology</td><td>Detectores personalizados</td><td>não digitalizados</td></tr><tr><td>Bomba</td><td>Harvard Apparatus</td><td>703602, 704402</td><td>Injetor e controlador Nanomite</td></tr><tr><td></td><td>Qualidade salina Biológico</td><td>114-055-721EA</td><td>Solução salina normal</td></tr><tr><td>Tesoura afiada de ponta romba</td><td>Braintree Scientific, Inc</td><td>SCT-S 508</td><td>Tesoura cirúrgica</td></tr><tr><td>Seringa</td><td>Covidien</td><td>22-257-150</td><td>Seringa de tuberculina de 1mL</td></tr><tr><td>TCS SP8 Microscópio espectral de mergulho</td><td>Leica</td><td>SP8</td><td>Pinça de microscópio de dois fótons</td></tr><tr><td>&nbsp;</td><td>Braintree Scientific, Inc</td><td>FC032</td><td>Fórceps de tecido</td></tr><tr><td>Xilazina&nbsp;</td><td>Anestésico</td> <td>VetOne</td><td>13985-704-10</td></tr></tbody></table>

microscopia subcelular intravital da glândula mamária

Intravital Subcellular Microscopy of the Mammary Gland

O leite é secretado a partir de células epiteliais alveolares polarizadas, que se diferenciam durante a gravidez dos brotos de uma extensa árvore ductular26 (Figura 2A). Os precursores da síntese do leite são assimilados através das membranas basais / laterais e os produtos completos são secretados através da superfície apical em um "espaço lácteo" central. O lado basal de cada alvéolo é coberto por uma matriz estrelada de células mioepiteliais (F...

Microscopia Subcelular Intravital da Glândula Mamária

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

Biologia

Lapso de tempo de imagem é utilizado para avaliar o comportamento dos principais pré-neoplásicas células epiteliais mamárias derivados de modelos de ratos geneticamente modificados de risco de câncer de mama para determinar se há correlação entre os parâmetros específicos de comportamento e distintas lesões genéticas.

Lapso de tempo de imagem de pré-neoplásicas primárias células epiteliais mamárias derivados de modelos de rato geneticamente modificado do cancro da mama

Microscopia intravital é uma ferramenta poderosa que permite a imagem latente vários processos biológicos em animais vivos. Neste artigo, é apresentado um método detalhado para a imagem dinâmica das estruturas subcelulares, tais como os grânulos secretores, nas glândulas salivares de ratos vivos.

Microscopia intravital para Estruturas de imagem Subcelulares em ratos vivos Expressando fluorescentes Proteínas

Um protocolo para a entrega intraductal não-invasivo de reagentes aquosas para a glândula mamaria do rato está descrito. O método aproveita injecção localizada nos mamilos das glândulas mamárias destinadas especificamente ductos mamários. Esta técnica é adaptável para uma variedade de compostos, incluindo o siRNA, agentes quimioterapêuticos e moléculas pequenas.

Injeção Intraductal para localizada Drug Delivery ao Rato glândula mamária

O protocolo de imunofluorescência indirecta descrito neste artigo permite a detecção e a localização de proteínas na glândula mamaria do rato. Um método completo é dada para preparar amostras da glândula mamaria, a efectuar imuno-histoquímica, para as secções de tecido imagem por microscopia de fluorescência, e para reconstruir imagens.

Imunofluorescência Indireta em cortes congelados de Rato glândula mamária

Este protocolo descreve o uso de microscopia multifotônica para executar imagiologia prolongado de lapso de tempo de interacções multicelulares em tempo real, in vivo, a resolução de células simples.

Estendida Time-lapse Imagem intravital do Real-time multicelular Dynamics no microambiente tumoral

Aqui, descrevemos uma técnica para a entrega localizada de reagentes na glândula mamária de coelho por meio de uma injeção intraductal. Além disso, descrevemos um protocolo para visualização e confirmação da entrega por ultrassonografia de alta resolução de agentes de contraste.

Entrega Intraductal ao Coelho glândula mamária

Aqui, apresentamos um protocolo para a isolação do rato inteiro, intacto, glândulas mamárias para investigar a expressão da matriz extracelular (ECM) e morfologia ductal. Mouse #4 glândulas abdominais foram extraídas dos ratos de 8-10 semanas nulíparas fêmeas, fixados em formalina tamponada neutra, seccionado e manchado usando imuno-histoquímica para proteínas de ECM.

Isolamento de glândulas mamárias de Mouse intacto, todo para a análise da expressão de matriz extracelular e morfologia da glândula

Uma abordagem é aqui apresentada para imagens intravitais de longo prazo usando janelas de silicone opticamente claras que podem ser coladas diretamente ao tecido/órgão de interesse e à pele. Essas janelas são mais baratas e mais versáteis do que outras atualmente utilizadas no campo, e a inserção cirúrgica causa inflamação e angústia limitadas aos animais.

Imagem intravital longitudinal através de janelas de silicone claras

Este protocolo descreve uma nova janela de imagem mamária com uma tampa substituível (R.MIW). A microscopia intravital após a implantação do R.MIW permite imagens longitudinais e de vários dias da glândula mamária saudável e doente com resolução celular durante os diferentes estágios de desenvolvimento.

Microscopia intravital longitudinal usando uma janela de imagem mamária com tampa substituível

O método de imagem intravital descrito aqui utiliza colágeno segunda geração harmônica e fluorescência endógena do cofator metabólico NAD(P)H para segmentar não invasivamente um microambiente tumoral não rotulado em compartimentos tumorais, estromaéreos e vasculares para análise aprofundada de imagens intravital 4D.

Uma abordagem de segmentação sem rótulos para imagens intravitais do microambiente tumoral mamário

O presente protocolo descreve a injeção intraductal de vetores virais via teto para entregar genes de interesse nas células epiteliais mamárias.

In vivo Entrega de Genes em Células Epiteliais Mamárias de Camundongos Através de Injeção Intraductal Mamária

Aprenda microscopia confocal e multiphoton intravital da glândula mamária lactante em camundongos. Este método permite visualizar dinâmica de gotículas lipídicas e morfologia glandular em tempo real.

Microscopia Subcelular Intravital da Glândula Mamária

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