この作品はアメリカの小児外科協会財団賞（AG）と健康K08DK098271（AG）の国立研究所メートがサポートされていました。

ウィスコンシン大学の動物実験委員会は、すべての手順を承認しました。 ソリューションの調製<ol><li>解剖緩衝液およびリンス液として1×リン酸緩衝生理食塩水（PBS）を使用します。 </li><li>ボトルにスクロースおよび場所の30グラムを計量することにより、30％スクロースを準備します。 1×PBS 99 mlを加え、1mlの10％アジ化ナトリウムを追加します。ショ糖の全てが溶解するまで十分に混合します。 4℃で保存、必要になるまで。 </li><li> 1×PBS、3％ウシ血清アルブミン（BSA）および0.1％トリトンX-100を混合して溶液を遮断準備。完全に混合し、必要になるまで冷蔵庫で保管してください。 </li><li> 1×PBS中にPFAの適切な量を秤量することによって1×PBS中の8％パラホルムアルデヒド（PFA）溶液を調製し、それが完全に溶解するまで65℃でインキュベートします。 -20℃の冷凍庫で25ミリリットルの分量を保存が必要になるまで。使用日の1×PBS中の4％PFAに希釈します。 </li></ol> 2.胚とガット解剖イオン<ol><li>制度動物実験委員会によると、タイミング妊娠マウスを安楽死させるし、PBS 1X氷冷を含む60ミリメートルペトリ皿に子宮を転送プロトコルを承認しました。 </li><li>鋭いハサミを使用して、解剖顕微鏡下で、胚を露出させるオープン子宮壁をカット。子宮から胚を除去し、PBS 1X氷冷を含むクリーンな60ミリメートルペトリ皿に置きます。 </li><li>氷冷1×PBS中で断頭により各胚を安楽死させます。あなたは、蛍光タンパク質を含むトランスジェニックマウスを使用している場合は、蛍光照明の下で、正のトランスジェニック胚を識別します。 </li><li>解剖顕微鏡を使用して、各胚から胃腸管を解剖。微細鉗子の使用、左側が上向きにされ、右側には、ペトリ皿の底に反するような胚を方向付けます。内臓を露出させるために胚から上半身の壁を削除します。背側体壁と内臓の間に微細な鉗子を挿入します。クロム胚から内臓を除去するために、はさみ状の切断動作で互いに対して鉗子をOSS。 </li><li>さらに離れて周囲の器官から胃腸管をサブ解剖した後、氷冷1×PBSを含む1.5 mlマイクロチューブに各GI管を配置します。 </li></ol> GI地帯の3固定<ol><li>氷冷1×PBSで各GI管を3回すすぎ、次いで4％PFAと交換してください。 1.5時間室温でロッキングプラットフォーム上で1.5mlの微小管における消化管を修正しました。 RTで消化管を5分間3回すすぎ、その後、ロッキングプラットフォーム上で1時間。この段階で、必要になるまで、0.1％アジ化ナトリウムを含有する1×PBS中30％スクロースに4℃でGIトラクトを格納します。 注：別の方法として、組織の完全性に影響することなく、1年まで、30％スクロース中の胚を格納します。 30％スクロース中のサンプルの保管は、免疫染色のために、またはクライオsectioniのため10月にいずれかのサンプルの後の処理を可能にしますNG。あるいは、以下に詳述免疫染色プロトコルを進めます。 </li></ol> 4.免疫染色プロトコル<ol><li>サンプルは30％スクロース中で保存されている場合は、ロッキングプラットフォーム上で、それらを1×PBSで20分間3回洗浄します。 </li><li> RTで1時間のためのロッキングプラットフォーム上でブロッキング溶液中に消化管を配置します。 </li><li>ブロッキング溶液を除去し、ロッキングプラットフォーム上で4℃でRTまたはO / Nで4時間のいずれかのためのブロッキング溶液中で希釈した一次抗体の適切な量の消化管をインキュベートします。 （患者から得られた血清）ヒト抗胡抗体1000希釈、1：ニワトリ抗緑色蛍光タンパク質（GFP）抗体1000希釈および1：ヤギ抗ChATの抗体の100倍希釈1を使用してください。 注：私たちは、患者から局所的に得られたヒト抗胡抗体を利用したが、抗胡抗体は、例えば、市販されており、マウス抗胡、（1での使用：500希釈）。 </li><li> 1×PBS中の消化管をすすぎます3回5分間、その後、ロッキングプラットフォーム上で室温で1時間。 </li><li> 4℃でのRTまたはO / Nで4時間のいずれかのためのロッキングプラットフォーム上のブロッキング溶液中で1：500に希釈した二次抗体を用いて1×PBSを交換してください。このようなDylight、ロバ抗ニワトリCy2とし、ロバ抗ヤギCy5のようなロバ抗ヒトアミン反応性染料を使用してください。ロバ抗マウスアミン反応性染料405：500希釈のマウス抗胡抗体を用いた場合は、1を使用します。 注：内因性GFPとtdTomato発現および発達年齢とともに増加して免疫染色を明確にする強さ。我々は、一般的に使用される抗体の量を低減し、また、組織の効率的な染色を確実にするために、染色溶液の150μlを、0.2 mlチューブ中に個別に胚勇気を免疫染色します。 </li><li> 1×PBS中の消化管に5分間、3回洗浄し、その後、ロッキングプラットフォーム上で室温で1時間。 </li><li>スライドガラス上にDAPIで蛍光封入剤を数滴を置きます。 GI TRAを浸します蛍光マウンティング培地中にCTと組織の上に直接カバーガラスを追加します。媒体-Gを取り付ける蛍光は半永久的なシールを提供する水溶性の化合物です。 注：このようなベクタシールドなどを使用し、蛍光封入フェージングおよび抗体の光退色防止媒体を装着するベースグリセロールです。これらの製品はいずれも、組織は4℃で長期間保存することを可能にします。 </li><li>共焦点顕微鏡を用いて蛍光体のそれぞれの画像をキャプチャします。 表3に示す蛍光団および使用するフィルタの励起波長を使用してください。 </li><li>画像をキャプチャするために使用される共焦点の種類に応じて、適切なソフトウェアを使用して、コンピュータ支援画像解析を実行します。 </li></ol>

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The authors have nothing to disclose.

私たちの研究室等はHSCR腸内の欠陥があっても神経節の小腸5,15,16に、無神経節結腸に限らず、近位に延長されていないことを示しています。これらの変化はENS神経密度と神経伝達物質表現型の変化を含み、HSCRを有する患者において観察された運動障害を説明することができます。私たちは、ENS形成の決定要因を理解するための努力で上記の技術を利用しています。具体的には、これ?...

運動、栄養吸収、およびローカルの血流を制御する機能腸管神経系（ENS）は、生命1に不可欠です。 ENSは、それらはニューロンおよびグリア細胞を含む神経節に分化腸を移動し、コロニー形成、増殖、神経堤細胞（NCC）によって形成されます。ヒルシュスプルング病（HSCR、男性でのオンラインメンデル遺伝）、1 4,000で出生の発生率とmultigeneic先天性障害は、破壊されたENSの形成を研究するためのプロトタイプの病気と考えることができます。 HSCRでは、NCCはに移動し、遠位後腸2の変数の長さを植民地化することができません。また、他の一般的な胃腸（GI）は、肛門直腸奇形、腸atresias、および運動障害などの小児集団における発達障害は、基本的なENS機能の障害と関連しており、おそらく微妙な、過小評価、解剖学的変化とで機能的変化と関連していますENS 3-6。そのため、私たちはENS形成の発達決定要因を理解することを可能にする技術は、病因および小児消化管障害の治療の可能性に光を当てることができます。 移行と植民地化の後、NCCは、それらの神経伝達物質の表現型に特異的なマーカーでニューロンに分化します。コリン作動性ニューロンは、腸溶性ニューロン7の約60％を含み、かつコリンアセチルトランスフェラーゼ（チャット）、興奮性神経伝達物質アセチルコリンのための合成酵素を染色することによって検出することができます。歴史的には、コリン作動性ニューロンを可視化する試みは、中枢神経系に対して向けられた抗体の抗原特異性（CNS）は、末梢神経系（PNS）のChAT 8-10に対してチャット異なるによって混乱しました。しかし、胎盤のChATに対する抗体は、両方の中央と周辺のChAT 11-13を認識し、我々は最近、visuaを可能にする技術が記載されています高感度のENSのコリン作動性ニューロンのlization以前の開発でチャットレポーター線14で達成されたよりも。 ここでは、ニューロンにおけるENSの神経伝達物質の発現を可視化するマウス胚GI管の、固定および免疫染色を解剖するための手法を提示します。これらの研究のために、我々は、 チャット-Creマウスを利用しているR26Rと交配：floxSTOP：チャットのCreを生成するためにtdTomato動物を、R26R：floxSTOP：tdTomatoマウス （原稿全体のChAT-CreをtdTomatoと定義されます）。これらの動物は、その後のChAT発現14を検出する蛍光レポーターの両方を発現するマウスを得るために、ホモ接合のChAT-GFPレポーターマウスと交配させました。これら二つのレポーター動物は、C57BL / 6Jバックグラウンド上にあり、（ジャクソン·ラボラトリーズ、バーハーバー、ME）は、市販されています。

<table><tbody><tr><td>Phosphate buffered saline</td><td>Oxoid</td><td>BR0014G</td><td></td></tr><tr><td>Sucrose</td><td>Fisher</td><td>S2</td><td></td></tr><tr><td>Sodium azide</td><td>Fisher</td><td>BP9221</td><td></td></tr><tr><td>Bovine serum albumin</td><td>Fisher</td><td>BP1605</td><td></td></tr><tr><td>Triton X-100</td><td>Sigma</td><td>X100</td><td></td></tr><tr><td>Paraformaldehyde</td><td>Sigma</td><td>158127</td><td></td></tr><tr><td>60 mm Petri dishes</td><td>Fisher</td><td>FB0875713A</td><td></td></tr><tr><td>Fluorescence microscope</td><td>Nikon</td><td>SMZ-18 stereoscope</td><td></td></tr><tr><td>Dissection microscope</td><td>Nikon</td><td>SMZ-18 stereoscope</td><td></td></tr><tr><td>Fine forceps</td><td>Fine science tools</td><td>11252-20</td><td></td></tr><tr><td>1.5 ml Eppendorf tubes</td><td>VWR</td><td>20170-038</td><td></td></tr><tr><td>Fluoromount-G</td><td>SouthernBiotech, Birmingham, AL</td><td>0100-01</td><td></td></tr><tr><td>Glass slides</td><td>Fisher</td><td>12-550-15</td><td></td></tr><tr><td>Cover glass</td><td>VWR</td><td>16004-330</td><td></td></tr><tr><td>Confocal microscope</td><td>Nikon</td><td>Nikon A1</td><td></td></tr><tr><td>Nikon Elements</td><td>Nikon</td><td></td><td></td></tr></tbody></table>

immunostaining to visualize murine enteric nervous system development

The enteric nervous system is formed by neural crest cells that proliferate, migrate and colonize the gut. Following colonization, neural crest cells must then differentiate into neurons with markers specific for their neurotransmitter phenotype. Cholinergic neurons, a major neurotransmitter phenotype in the enteric nervous system, are identified by staining for choline acetyltransferase (ChAT), the synthesizing enzyme for acetylcholine. Historical efforts to visualize cholinergic neurons have been hampered by antibodies with differing specificities to central nervous system versus peripheral nervous system ChAT. We and others have overcome this limitation by using an antibody against placental ChAT, which recognizes both central and peripheral ChAT, to successfully visualize embryonic enteric cholinergic neurons. Additionally, we have compared this antibody to genetic reporters for ChAT and shown that the antibody is more reliable during embryogenesis. This protocol describes a technique for dissecting, fixing and immunostaining of the murine embryonic gastrointestinal tract to visualize enteric nervous system neurotransmitter expression.

我々は以前のChAT発現14を検出GFPおよびtdTomato蛍光レポーターの両方を発現するマウスの生成を記載しています。簡単に説明すると、 チャットCreマウスは、R26Rと交配させた：floxSTOP：チャットのCreを生成するためにtdTomato動物; R26R：floxSTOP：（チャット-CreをtdTomatoと呼ばれる）tdTomatoマウスを。これらの動物は、その後、ホモ接合のChAT-GFPレ...

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マウスの腸管神経系の発達を可視化するための免疫染色 |発生生物学 |ビデオ

Immunostaining to Visualize Murine Enteric Nervous System Development

The enteric nervous system is formed by neural crest cells that proliferate, migrate and colonize the gut. Neural crest cells differentiate into neurons with markers specific for their neurotransmitter phenotype. This protocol describes a technique for dissecting, fixing and immunostaining of the murine embryonic gastrointestinal tract to visualize enteric nervous system neurotransmitter expression.

マウス腸神経系の開発を可視化するために免疫染色

The enteric nervous system is formed by neural crest cells that proliferate, migrate and colonize the gut. Following colonization, neural crest cells must ...

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11.3K ビュー.  University of Wisconsin School of Medicine and Public Health. The enteric nervous system is formed by neural crest cells that proliferate, migrate and colonize the gut. Neural crest cells differentiate into neurons with markers specific for their neurotransmitter phenotype. This protocol describes a technique for dissecting, fixing and immunostaining of the murine embryonic gastrointestinal tract to visualize enteric nervous system neurotransmitter expression.

ビデオ: Immunostaining to Visualize Murine Enteric Nervous System Development

Multimodal Imaging of Stem Cell Implantation in the Central Nervous System of Mice

During the past decade, stem cell transplantation has gained increasing interest as primary or secondary therapeutic modality for a variety of diseases, both in preclinical and clinical studies. However, to date results regarding functional outcome and/or tissue regeneration following stem cell transplantation are quite diverse. Generally, a clinical benefit is observed without profound understanding of the underlying mechanism(s)1. Therefore, multiple efforts have led to the development of different molecular imaging modalities to monitor stem cell grafting with the ultimate aim to accurately evaluate survival, fate and physiology of grafted stem cells and/or their micro-environment. Changes observed in one or more parameters determined by molecular imaging might be related to the observed clinical effect. In this context, our studies focus on the combined use of bioluminescence imaging (BLI), magnetic resonance imaging (MRI) and histological analysis to evaluate stem cell grafting.
BLI is commonly used to non-invasively perform cell tracking and monitor cell survival in time following transplantation2-7, based on a biochemical reaction where cells expressing the Luciferase-reporter gene are able to emit light following interaction with its substrate (e.g. D-luciferin)8, 9. MRI on the other hand is a non-invasive technique which is clinically applicable10 and can be used to precisely locate cellular grafts with very high resolution11-15, although its sensitivity highly depends on the contrast generated after cell labeling with an MRI contrast agent. Finally, post-mortem histological analysis is the method of choice to validate research results obtained with non-invasive techniques with highest resolution and sensitivity. Moreover end-point histological analysis allows us to perform detailed phenotypic analysis of grafted cells and/or the surrounding tissue, based on the use of fluorescent reporter proteins and/or direct cell labeling with specific antibodies.
In summary, we here visually demonstrate the complementarities of BLI, MRI and histology to unravel different stem cell- and/or environment-associated characteristics following stem cell grafting in the CNS of mice. As an example, bone marrow-derived stromal cells, genetically engineered to express the enhanced Green Fluorescent Protein (eGFP) and firefly Luciferase (fLuc), and labeled with blue fluorescent micron-sized iron oxide particles (MPIOs), will be grafted in the CNS of immune-competent mice and outcome will be monitored by BLI, MRI and histology (Figure 1).

This article describes an optimized sequence of events for multimodal imaging of cellular grafts in rodent brain using: (i) in vivo bioluminescence and magnetic resonance imaging, and (ii) post mortem histological analysis. Combining these imaging modalities on a single animal allows cellular graft evaluation with high resolution, sensitivity and specificity.

マウスの中枢神経系における幹細胞注入のマルチモーダルイメージング

During the past decade, stem cell transplantation has gained increasing interest as primary or secondary therapeutic modality for a variety of diseases, ...

神経科学

Whole Mount Immunofluorescent Staining of the Neonatal Mouse Retina to Investigate Angiogenesis In vivo

Angiogenesis is the complex process of new blood vessel formation defined by the sprouting of new blood vessels from a pre-existing vessel network. Angiogenesis plays a key role not only in normal development of organs and tissues, but also in many diseases in which blood vessel formation is dysregulated, such as cancer, blindness and ischemic diseases. In adult life, blood vessels are generally quiescent so angiogenesis is an important target for novel drug development to try and regulate new vessel formation specifically in disease. In order to better understand angiogenesis and to develop appropriate strategies to regulate it, models are required that accurately reflect the different biological steps that are involved. The mouse neonatal retina provides an excellent model of angiogenesis because arteries, veins and capillaries develop to form a vascular plexus during the first week after birth. This model also has the advantage of having a two-dimensional (2D) structure making analysis straightforward compared with the complex 3D anatomy of other vascular networks. By analyzing the retinal vascular plexus at different times after birth, it is possible to observe the various stages of angiogenesis under the microscope. This article demonstrates a straightforward procedure for analyzing the vasculature of a mouse retina using fluorescent staining with isolectin and vascular specific antibodies.

Whole Mount Immunofluorescent Staining of the Neonatal Mouse Retina to Investigate Angiogenesis In vivo

The neonatal murine retina provides a well characterized physiological model of angiogenesis, which permits investigations of the roles of different genes or drugs that modulate angiogenesis in an in vivo context. Immunofluorescent staining to accurately visualize the vascular plexus is pivotal to the success of these types of studies.

血管新生を調査するために新生児マウス網膜の全体のマウント免疫蛍光染色<em生体内で&gt;</em

Angiogenesis  is the complex process of new blood vessel formation defined by the sprouting  of new blood vessels from a pre-existing vessel network. ...

An In-vitro Preparation of Isolated Enteric Neurons and Glia from the Myenteric Plexus of the Adult Mouse

The enteric nervous system is a vast network of neurons and glia running the length of the gastrointestinal tract that functionally controls gastrointestinal motility. A procedure for the isolation and culture of a mixed population of neurons and glia from the myenteric plexus is described. The primary cultures can be maintained for over 7 days, with connections developing among the neurons and glia. The longitudinal muscle strip with the attached myenteric plexus is stripped from the underlying circular muscle of the mouse ileum or colon and subjected to enzymatic digestion. In sterile conditions, the isolated neuronal and glia population are preserved within the pellet following centrifugation and plated on coverslips. Within 24-48 hr, neurite outgrowth occurs and neurons can be identified by pan-neuronal markers. After two days in culture, isolated neurons fire action potentials as observed by patch clamp studies. Furthermore, enteric glia can also be identified by GFAP staining. A network of neurons and glia in close apposition forms within 5 - 7 days. Enteric neurons can be individually and directly studied using methods such as immunohistochemistry, electrophysiology, calcium imaging, and single-cell PCR. Furthermore, this procedure can be performed in genetically modified animals. This methodology is simple to perform and inexpensive. Overall, this protocol exposes the components of the enteric nervous system in an easily manipulated manner so that we may better discover the functionality of the ENS in normal and disease states.

An In-vitro Preparation of Isolated Enteric Neurons and Glia from the Myenteric Plexus of the Adult Mouse

We demonstrate a cell culture protocol for the direct study of neuronal and glial components of the enteric nervous system. A neuron/glia mixed culture on coverslips is prepared from the myenteric plexus of adult mouse providing the ability to examine individual neuron and glia function by electrophysiology, immunohistochemical, etc.

不定冠詞<em&gt;インビトロ</em成体マウスの筋層間神経叢から分離腸ニューロンとグリアの&gt;準備

The enteric nervous system is a vast network of neurons and glia running the length of the gastrointestinal tract that functionally controls ...

A Quantitative Cell Migration Assay for Murine Enteric Neural Progenitors

Neural crest cells (NCC) are a transient and multipotent cell population that originates from the dorsal neural tube and migrates extensively throughout the developing vertebrate embryo. In addition to providing peripheral glia and neurons, NCC generate melanocytes as well as most of the cranio-facial skeleton. NCC migration and differentiation is controlled by a combination of their axial origin along the neural tube and their exposure to regionally distinct extracellular cues. Such contribution of extracellular ligands is especially evident during the formation of the enteric nervous system (ENS), a complex interconnected network of neural ganglia that locally controls (among other things) gut muscle movement and intestinal motility. Most of the ENS is derived from a small initial pool of NCC that undertake a long journey in order to colonize - in a rostral to caudal fashion - the entire length of the prospective gut. Among several signaling pathways known to influence enteric NCC colonization, GDNF/RET signaling is recognized as the most important. Indeed, spatiotemporally controlled secretion of the RET ligand GDNF by the gut mesenchyme is chiefly responsible for the attraction and guidance of RET-expressing enteric NCC to and within the embryonic gut. Here, we describe an ex vivo cell migration assay, making use of a transgenic mouse line possessing fluorescently labeled NCC, which allows precise quantification of enteric NCC migration potential in the presence of various growth factors, including GDNF.

We present an ex vivo cell migration assay that allows precise quantification of enteric neural crest cell migration potential in the presence of various growth factors.

マウス腸の神経前駆細胞の定量細胞遊走アッセイ

Neural crest cells (NCC) are a transient and multipotent cell population that originates from the dorsal neural tube and migrates extensively throughout ...

In Situ Ca2+ Imaging of the Enteric Nervous System

Reflex behaviors of the intestine are controlled by the enteric nervous system (ENS). The ENS is an integrative network of neurons and glia in two ganglionated plexuses housed in the gut wall. Enteric neurons and enteric glia are the only cell types within the enteric ganglia. The activity of enteric neurons and glia is responsible for coordinating intestinal functions. This protocol describes methods for observing the activity of neurons and glia within the intact ENS by imaging intracellular calcium (Ca2+) transients with fluorescent indicator dyes. Our technical discussion focuses on methods for Ca2+ imaging in whole-mount preparations of the myenteric plexus from the rodent bowel. Bulk loading of ENS whole-mounts with a high-affinity Ca2+ indicator such as Fluo-4 permits measurements of Ca2+ responses in individual neurons or glial cells. These responses can be evoked repeatedly and reliably, which permits quantitative studies using pharmacological tools. Ca2+ responses in cells of the ENS are recorded using a fluorescence microscope equipped with a cooled charge-coupled device (CCD) camera. Fluorescence measurements obtained using Ca2+ imaging in whole-mount preparations offer a straightforward means of characterizing the mechanisms and potential functional consequences of Ca2+ responses in enteric neurons and glial cells.

In Situ Ca2+ Imaging of the Enteric Nervous System

The enteric nervous system (ENS) is a network of neurons and glia located in the gut wall that controls intestinal reflexes. This protocol describes methods for recording the activity of enteric neurons and glia in live preparations of ENS using Ca2+ imaging.

腸神経系のその場のCa 2+イメージングに

Reflex behaviors of the intestine are controlled by the enteric nervous system (ENS). The ENS is an integrative network of neurons and glia in two ...

感染と自然免疫を研究するためのマウス脳胚混合細胞培養

Establishing Mixed Neuronal and Glial Cell Cultures from Embryonic Mouse Brains to Study Infection and Innate Immunity

Models of the central nervous system (CNS) must recapitulate the complex network of interconnected cells found in vivo. The CNS consists primarily of neurons, astrocytes, oligodendrocytes, and microglia. Due to increasing efforts to replace and reduce animal use, a variety of in vitro cell culture systems have been developed to explore innate cell properties, which allow the development of therapeutics for CNS infections and pathologies. Whilst certain research questions can be addressed by human-based cell culture systems, such as (induced) pluripotent stem cells, working with human cells has its own limitations with regard to availability, costs, and ethics. Here, we describe a unique protocol for isolating and culturing cells from embryonic mouse brains. The resulting mixed neural cell cultures mimic several cell populations and interactions found in the brain in vivo. Compared to current equivalent methods, this protocol more closely mimics the characteristics of the brain and also garners more cells, thus allowing for more experimental conditions to be investigated from one pregnant mouse. Further, the protocol is relatively easy and highly reproducible. These cultures have been optimized for use at various scales, including 96-well based high throughput screens, 24-well microscopy analysis, and 6-well cultures for flow cytometry and reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis. This culture method is a powerful tool to investigate infection and immunity within the context of some of the complexity of the CNS with the convenience of in vitro methods.

著者スポットライト:感染と自然免疫を研究するためのマウス胚脳からの混合ニューロン細胞培養とグリア細胞培養の確立  

This protocol presents a unique way of generating central nervous system cell cultures from embryonic day 17 mouse brains for neuro(immuno)logy research. This model can be analyzed using various experimental techniques, including RT-qPCR, microscopy, ELISA, and flow cytometry.

感染と自然免疫を研究するためのマウス胚性脳からの混合神経細胞およびグリア細胞培養の確立

Models of the central nervous system (CNS) must recapitulate the complex network of interconnected cells found in vivo. The CNS consists primarily of ...

Using a Whole-mount Immunohistochemical Method to Study the Innervation of the Biliary Tract in Suncus murinus

This work describes the whole-mount immunohistochemistry staining method in detail, using neurofilament protein antibody to label the innervation of the biliary tract in Suncus murinus (S. murinus&#160;). First, the specimen was dissected from S. murinus and fixed in 4% paraformaldehyde (PFA). Second, an enzymatic treatment and potential endogenous peroxidase inactivation were performed. The specimen was then exposed to the primary antibody, anti-neurofilament protein antibody, for 3-6 days. It was then incubated with the secondary antibody conjugated with horseradish peroxidase. The color reaction was revealed by reacting the specimen with a 3,3&#39;-diaminobenzidine (DAB) substrate. This method can be applied to analyze the innervation of all visceral organs. Furthermore, this protocol can also be adapted to test other neuronal antibodies, but optimization of the antibodies should be done first. This method was originally introduced by Kuratani and Tanaka1,2,3.

Using a Whole-mount Immunohistochemical Method to Study the Innervation of the Biliary Tract in Suncus murinus

A whole-mount immunohistochemical approach, to visualize neurofilament protein expression in the extrahepatic biliary tract in Suncus murinus. is presented here. This protocol can be used to analyze the innervation of all visceral organs in S. murinus or other species.

全身免疫組織化学的方法を用いて胆道の内腔を調べる<em&gt; Suncus murinus</em

This work describes the whole-mount immunohistochemistry staining method in detail, using neurofilament protein antibody to label the innervation of the ...

生物学

Immunofluorescence Staining for Visualizing Proliferated Neural Stem Cells in Mouse Brain Sections

Source: Quintanilla, L. J., et al. <a href="https://app.jove.com/v/59237">Assaying Circuit Specific Regulation of Adult Hippocampal Neural Precursor Cells.</a> J. Vis. Exp. (2019)
This video demonstrates the procedure for staining newly proliferated neural stem cells in brain tissue sections using EdU incorporation, followed by immunofluorescence to visualize specific proteins, enhancing the understanding of cell dynamics and proliferation.

マウス脳切片における増殖神経幹細胞の可視化のための免疫蛍光染色

All procedures involving animal samples have been reviewed and approved by the appropriate animal ethical review committee.
1. Immunohistochemistry

	...

マウス腸神経系の開発を可視化するために免疫染色

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マウスの中枢神経系における幹細胞注入のマルチモーダルイメージング

血管新生を調査するために新生児マウス網膜の全体のマウント免疫蛍光染色

不定冠詞

マウス腸の神経前駆細胞の定量細胞遊走アッセイ

腸神経系のその場のCa ²⁺イメージングに

感染と自然免疫を研究するためのマウス胚性脳からの混合神経細胞およびグリア細胞培養の確立

感染と自然免疫を研究するためのマウス胚性脳からの混合神経細胞およびグリア細胞培養の確立

感染と自然免疫を研究するためのマウス胚性脳からの混合神経細胞およびグリア細胞培養の確立

全身免疫組織化学的方法を用いて胆道の内腔を調べる

マウス脳切片における増殖神経幹細胞の可視化のための免疫蛍光染色