Name: murine superficial lymph node surgery
Uploaded: 2012-05-21T12:00:00
Description: Watch this Scientific Journal Video about Murine Superficial Lymph Node Surgery at JoVE.com

Vi takker Sylvie Lesage og alle lab medlemmer til kritisk læsning af protokollen. Dette arbejde blev støttet af en bevilling (MOP-77.545) fra den canadiske Institutes of Health Research (CIHR). Nathalie Labrecque understøttes af en FRSQ Senior Scholarship og Melissa Mathieu modtog en NSERC Alexander Graham Bell Canada Graduate Scholarship.

1. Forberedelse før operationen Mus blev behandlet i overensstemmelse med den canadiske Rådet om Animal Care retningslinjer. <ol><li> Anesthetise mus ved injektion af ketamin / xylazin (150/300 mg / kg, ip) eller ved inhalation af isofluran (2%, 1L oxygen). Hvis det er muligt, bør isofluran kan anvendes, da det giver mulighed for en bedre kontrol af anæstesi tid og dybde. Desuden er oxygen administreres på samme tidspunkt, som understøtter de fysiologiske funktioner af dyret. Desuden...

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Vi har beskrevet en protokol LN kirurgi, som kan anvendes til mange eksperimentelle systemer. Selv meget nyttigt at følge immunresponser eller immuncelle homeostase, denne teknik har nogle begrænsninger. For det første kan kun fire LN blive høstet. For det andet skal immunsystemet undersøgt respons være systemisk eller forekommer i de overfladiske ingvinale eller brachiale (hud-drænende) LN. Endelig er antallet af høstede celler påvirkes af kvaliteten af ​​LN fjernelse, en teknisk begrænsning som kan bidra...

<table border="1"><tbody><tr><td> Reagens Navn </td><td> Firma </td><td> Katalog nr. </td><td> Kommentarer </td></tr><tr><td> Graefe fint mønster Premium Tang </td><td> Harvard Apparatus </td><td> 52-2144 </td><td> Stærkt Svejfet, 10 cm (4 i), 0,8 mm spids </td></tr><tr><td> Michel Clip påfører og fjerner Tang </td><td> Harvard Apparatus </td><td> 52-3779 </td><td> 12,5 cm (5 in) </td></tr><tr><td> Michel Clip 100 </td><td> Harvard Apparatus </td><td> 52-3746 </td><td> 7,5-1,75 mm </td></tr><tr><td> Temgesic (buprenorphinhydrochlorid) </td><td> Schering-Plough </td><td></td><td></td></tr><tr><td> Vetalar (ketamin-hydrochlorid) </td><td> Boniche Animal Healthcare </td><td></td><td> DIN: 01989529 </td></tr><tr><td> Rompun (xylazin) </td><td> Bayer </td> &lt;td&gt; </td><td> DIN: 02169592 </td></tr><tr><td> Isofluran </td><td> ABBOTT </td><td></td><td> DIN: 02032384 </td></tr><tr><td> Hypotears øjensalve </td><td> Novartis Ophtalmics </td><td></td><td> DIN: 02133288 </td></tr><tr><td> Baxedin (Clorhexidine gluconat 2% i isopropylalkohol 70%) </td><td> Omega </td><td> L0000017 </td><td> DIN: 02251477 </td></tr></tbody></table>

murine superficial lymph node surgery

Inden for immunologi, til at forstå progressionen af ​​en immunrespons mod en vaccine, en infektion eller en tumor, bliver respons ofte fulgt over tid. Tilsvarende studiet af lymfocyt homeostase kræver tidsforloebet eksperimenter. Udførelse af disse forsøg inden for den samme mus er ideel til at reducere den eksperimentelle variation samt antallet af anvendte mus. Blod tilbagetrækning tillader udførelse af tidsforløbet eksperimenter, men kun giver oplysning om cirkulerende lymfocytter og tilvejebringer et begrænset antal celler 1-4. Siden lymfocytter cirkulerer gennem kroppen og er bosat i lymfeknuderne har forskellige egenskaber, er det vigtigt at undersøge begge steder. Sekventiel fjernelse af lymfeknuder ved operation tilvejebringer en unik mulighed for at følge en immunreaktion eller immun-celleekspansion in samme mus over tid. Desuden er denne teknik giver mellem 1-2x10 6 celler pr lymfeknude, der er tilstrækkelig til at perform fænotypisk karakterisering og / eller funktionelle assays. Sekventiel lymfeknude operation eller lymphadenectomy held har været anvendt af os og andre 5-11. Her beskriver vi, hvordan brachial og inguinale lymfeknuder kan fjernes ved at gøre en lille indsnit i huden på en bedøvet mus. Da operationen er overfladisk og gjort hurtigt, musen genvinder meget hurtigt, heler godt, og ikke oplever store smerter. Hver anden dag, er det muligt at høste en eller to lymfeknuder muliggør tidsforløbet eksperimenter. Denne teknik er således egnet til at studere egenskaber lymfeknude-bosiddende lymfocytter over tid. Denne fremgangsmåde er egnet til forskellige eksperimentelle designs, og vi mener, at mange laboratorier vil have gavn af at udføre sekventielle lymfeknude operationer.

Watch this Scientific Journal Video about Murine Superficial Lymph Node Surgery at JoVE.com

Murine Superficial Lymph Node Surgery

At følge progressionen af ​​en immunreaktion over tid i samme mus, kan lymfeknuder sekventielt fjernes ved kirurgi. Her beskriver vi, hvordan denne teknik kan udføres.

Murin Overfladisk lymfeknude Kirurgi

In the field of immunology, to understand the progression of an immune response against a vaccine, an infection or a tumour, the response is often ...

Research

JoVE Journal

Immunology and Infection

Intravital Microscopy of the Inguinal Lymph Node

Lymph nodes (LN's), located throughout the body, are an integral component of the immune system. They serve as a site for induction of adaptive immune ...

Ex vivo Imaging of T Cells in Murine Lymph Node Slices with Widefield and Confocal Microscopes

Ex vivo Imaging of T Cells in Murine Lymph Node Slices with Widefield and Confocal Microscopes

Na&iuml;ve T cells continuously traffic to secondary lymphoid organs, including peripheral lymph nodes, to detect rare expressed antigens. The migration ...

Intravital Imaging of the Mouse Popliteal Lymph Node

Intravital Billeddannelse af mus popliteale lymfeknude

Lymph nodes (LNs) are secondary lymphoid organs, which are strategically located throughout the body to allow for trapping and presentation of foreign ...

Generation of Lymph Node-fat Pad Chimeras for the Study of Lymph Node Stromal Cell Origin

Generering af lymfeknuder-fedt Pad kimærer for Studiet af lymfeknuder stromacelle Origin

The stroma is a key component of the lymph node structure and function. However, little is known about its origin, exact cellular composition and the ...

Isolation of Murine Lymph Node Stromal Cells

Isolering af murine lymfeknuder stromacellers

Secondary lymphoid organs including lymph nodes are composed of stromal cells that provide a structural environment for homeostasis, activation and ...

Myeloid Cell Isolation from Mouse Skin and Draining Lymph Node Following Intradermal Immunization with Live Attenuated Plasmodium Sporozoites

Myeloid Cell Isolation from Mouse Skin and Draining Lymph Node Following Intradermal Immunization with Live Attenuated Plasmodium Sporozoites

Myeloid Cell Isolation fra Mouse Hud og drænende lymfeknude Efter Intrakutan Vaccination med levende svækket<em&gt; Plasmodium</em&gt; sporozoitter

Malaria infection begins when the sporozoite stage of Plasmodium is inoculated into the skin of a mammalian host through a mosquito bite. The highly ...

A CFSE-based Assay to Study the Migration of Murine Skin Dendritic Cells into Draining Lymph Nodes During Infection with Mycobacterium bovis Bacille Calmette-Gu&#233;rin

A CFSE-based Assay to Study the Migration of Murine Skin Dendritic Cells into Draining Lymph Nodes During Infection with Mycobacterium bovis Bacille Calmette-GuÃƒÆ’Ã†'Ãƒâ€šÃ‚Â©rin

En CFSE-baserede Assay til at studere den Migration af murine Skin dendritiske celler i drænende lymfeknuder Under Infektion med<em&gt; Mycobacterium bovis</em&gt; Bacille Calmette-Guérin

Dendritic cells (DCs) are important for initiating immune responses, in part through their ability to acquire and shuttle antigen to the draining lymph ...

Murine Full-thickness Skin Transplantation

Murint Full-tykkelse hud Transplantation

Murine full-thickness skin transplantation is a well-established in vivo model to study alloimmune response and graft rejection. Despite its limited ...

Intracellular Staining and Flow Cytometry to Identify Lymphocyte Subsets within Murine Aorta, Kidney and Lymph Nodes in a Model of Hypertension

Intracellulær Farvning og flowcytometri at identificere lymfocytundergrupper inden Murine Aorta, Nyre og lymfeknuder i en model af hypertension

It is now well known that T lymphocytes play a critical role in the development of several cardiovascular diseases1,2,3,4,5. For example, studies from our ...

Collection and Processing of Lymph Nodes from Large Animals for RNA Analysis: Preparing for Lymph Node Transcriptomic Studies of Large Animal Species

Indsamling og behandling af lymfeknuder fra store dyr til RNA analyse: forberede lymfeknude transkriptom undersøgelser af store dyrearter

Large animals (both livestock and wildlife) serve as important reservoirs of zoonotic pathogens, including Brucella, Mycobacterium bovis, Salmonella, and ...