Name: transurethral induction of mouse urinary tract infection
Uploaded: 2010-08-05T17:30:00
Description: Watch this Scientific Journal Video about Transurethral Induction of Mouse Urinary Tract Infection at JoVE.com

Die Autoren bedanken sich bei Shelly Xie und Jennifer Payne für ihre technische Unterstützung durch Experten bestätigt. Diese Arbeit wurde ermöglicht durch Zuschüsse aus dem Society of Pediatric Urology und der Stanford Pediatric Research Fund.

I. Vorbereitung der Harnröhren-Katheter <ol><li> Für jede Maus Blase und Niere entnommen werden, statt 5-6 Edelstahlkugeln (Blase, 1,6 mm Größe oder 0,9-2,0 mm Blend) oder Zirkonoxid-Perlen (Niere, 0,5 mm Größe) in einem Kryoröhrchen Rohr bzw.. Autoclave die Perle enthaltenden Röhrchen. </li><li> Nach der Rohre auf Umgebungstemperatur abkühlen, 200 Mikroliter steriles PBS zu jeder Edelstahl bead-haltigen Schlauch und 400 Mikroliter steriles PBS in jedes Zirkonoxid bead-haltigen Röhre. </li><li> Wir bereiten ...

<ol>
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Die Induktion der Maus Harnwegsinfektion durch transurethrale Inokulation von Bakterien ist eine seit langem etablierte, aber technisch anspruchsvollen Verfahren 11. Hung und Hultgren kürzlich veröffentlichten einen ausgezeichneten Überblick über transurethralen Techniken der Maus Harnwegsinfektion 13. In diesem Papier versuchen wir, weiter zu veranschaulichen, die Feinheiten dieser experimentelle Ansatz. Anfänger können ihre Technik, indem Sie eine Mittellinie La...

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		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>Cryovial tubes, 2.0 ml polypropylene (no caps)</td>
<td>&nbsp;</td>
<td>E&amp;K Scientific</td>
<td>640201</td>
<td>&nbsp;</td>
<tr>
<td>Caps for cryovial tubes, with 'O' ring</td>
<td>&nbsp;</td>
<td>E&amp;K Scientific</td>
<td>440011-N</td>
<td>&nbsp;</td>
<tr>
<td>Bullet Blender</td>
<td>&nbsp;</td>
<td> Next Advance</td>
<td>BBX24</td>
<td>&nbsp;</td>
<tr>
<td>Zirconium Oxide Beads (0.5mm)</td>
<td>&nbsp;</td>
<td>Next Advance</td>
<td>ZROB05</td>
<td>&nbsp;</td>
<tr>
<td>Stainless Steel Beads (1.6mm)</td>
<td>&nbsp;</td>
<td>Next Advance</td>
<td>SSB16</td>
<td>&nbsp;</td>
<tr>
<td>Stainless Steel Bead Blend (0.9-2.0mm)</td>
<td>&nbsp;</td>
<td>Next Advance</td>
<td>SSB14B</td>
<td>&nbsp;</td>
<tr>
<td>CBA/J Mice, 8-12 week old female</td>
<td>&nbsp;</td>
<td>Jackson Laboratories</td>
<td>000656</td>
<td>&nbsp;</td>
<tr>
<td>Phosphate Buffered Saline (NaCl 137mM, KCl 2.7mM, Na2HPO4 4.3mM, KH2PO4 1.4mM)</td>
<td>&nbsp;</td>
<td>EMD</td>
<td>EM-SX0420-13 (NaCl) EM-PX1405-1 (KCl) EM1.06585.0 (Na2HPO4) EM-5108-1 (KH2PO4)</td>
<td>&nbsp;</td>
<tr>
<td>LB agar</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>LB broth</td>
<td>&nbsp;</td>
<td>EMD</td>
<td>1.10285.0500</td>
<td>&nbsp;</td>
<tr>
<td>Isoflurane (Aerrane)</td>
<td>&nbsp;</td>
<td>Baxter</td>
<td>NDC 10019-773-60</td>
<td>&nbsp;</td>
<tr>
<td>Inoculating loops</td>
<td>&nbsp;</td>
<td>Fisher Scientific</td>
<td>22-363-601</td>
<td>&nbsp;</td>
<tr>
<td>Eosin-methylene blue (EMB) agar powder</td>
<td>&nbsp;</td>
<td>Himedia Laboratories</td>
<td>MM022-500G</td>
<td>&nbsp;</td>
<tr>
<td>MacConkey agar powder</td>
<td>&nbsp;</td>
<td>Difco</td>
<td>212123</td>
<td>&nbsp;</td>
<tr>
<td>LB agar powder</td>
<td>&nbsp;</td>
<td>Difco</td>
<td>244520</td>
<td>&nbsp;</td>
<tr>
<td>Intramedic non-radiopaque polyethylene tubing, PE10 [inner dimension 0.28 mm (0.011 inch); outer dimension 0.61 mm (0.024 inch)]</td>
<td>&nbsp;</td>
<td>BD Biosciences</td>
<td>427401</td>
<td>&nbsp;</td>
<tr>
<td>30 gauge needles, 1/2 inch</td>
<td>&nbsp;</td>
<td>BD Biosciences</td>
<td>305106</td>
<td>&nbsp;</td>
<tr>
<td>1 ml tuberculin Slip-Tip syringe</td>
<td>&nbsp;</td>
<td>BD Biosciences</td>
<td>309602</td>
<td>&nbsp;</td>
<tr>
<td>3 ml Luer-Lock syringe</td>
<td>&nbsp;</td>
<td>BD Biosciences</td>
<td>309585</td>
<td>&nbsp;</td>		</tbody>
		</table>
		</div>

transurethral induction of mouse urinary tract infection

Uropathogene Bakterienstämme von Interesse sind, auf Agar gewachsen. Generell uropathogene<em&gt; E. coli</em&gt; (UPEC) und andere Stämme können über Nacht auf Luria-Bertani (LB)-Agar bei 37 ° C in der Umgebungsluft. UPEC Stämme wachsen als gelblich-weiße durchscheinende Kolonien auf LB-Agar. Nach der Bestätigung der entsprechenden Koloniemorphologie sind einzelne Kolonien griff dann in Brühe gezüchtet werden. LB-Medium kann für die meisten uropathogene Bakterienstämmen verwendet werden können. Zwei serielle, über Nacht LB Bouillon-Kulturen eingesetzt, um die Expression von Typ-I-Pili, eine gut definierte Virulenzfaktor für uropathogene Bakterien zu verbessern. Bouillon-Kulturen sind auf die gewünschte Konzentration in phosphatgepufferter Salzlösung (PBS). Acht bis 12 Wochen alte weibliche Mäuse werden unter Isofluran-Narkose gelegt und transurethral mit Bakterien mit Polyethylen-Rohr bedeckt 30-Gauge-Spritzen geimpft. Typische Inokula, die empirisch für jede bakterielle / Maus-Stamm-Kombination ermittelt werden, sind 10<sup&gt; 6</sup&gt; Bis 10<sup&gt; 8</sup&gt; KBE pro Maus in 10 bis 50 Mikroliter PBS. Nachdem die gewünschte Infektion Zeitraum (ein Tag bis zu mehreren Wochen), Urinproben und der Blase und beide Nieren entnommen. Jedes Organ ist Hackfleisch, platziert in PBS und homogenisiert in einem Blue Bullet Homogenisator. Urin-und Gewebehomogenaten werden seriell in PBS verdünnt und über geeignete Agar. Am nächsten Tag sind koloniebildende Einheiten gezählt.

Watch this Scientific Journal Video about Transurethral Induction of Mouse Urinary Tract Infection at JoVE.com

Transurethral Induction of Mouse Urinary Tract Infection

Dieses Video demonstriert Methoden zur transurethral induzieren Maus Harnwegsinfektionen und zu quantifizieren, das Ausmaß der daraus resultierenden Infektionen.

Transurethrale Induktion von Maus Harnwegsinfekt

Uropathogenic bacterial strains of interest are grown on agar. Generally, uropathogenic E. coli (UPEC) and other strains can be grown overnight on ...

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