Name: Verzameling van neusspoelingsvloeistof bij muizen zonder bloedverontreiniging (Video) | JoVE Journal
Uploaded: 2025-07-11T15:10:00.000+00:00
Duration: 5 min 12 s
Description: Leer een methode voor het verzamelen van neusspoelingsvloeistof bij muizen zonder bloedverontreiniging. Deze techniek verbetert de nauwkeurigheid van immunologische analyses en detectie van luchtweginfecties.

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<li>Park, A. M., Tsunoda, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Forensic+luminol+reaction+for+detecting+fecal+occult+blood+in+experimental+mice%5BTitle%5D)+AND+%22Biotechniques%22%5BJournal%5D)">Forensic luminol reaction for detecting fecal occult blood in experimental mice.</a> Biotechniques. 65 (4), 227-230 (2018).</li>
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<li>Castigli, E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Impaired+IgA+class+switching+in+APRIL-deficient+mice%5BTitle%5D)+AND+%22Proc+Natl+Acad+Sci+U+S+A%22%5BJournal%5D)">Impaired IgA class switching in APRIL-deficient mice.</a> Proc Natl Acad Sci U S A. 101 (11), 3903-3908 (2004).</li>
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</ol>

<table><tbody><tr><td>Animal fixation needle</td><td>Natsume Seisakusho Co., Ltd., Tokyo, Japan</td><td>KN-358-A</td><td>We fixed a mouse with animal fixation needles. Stainless steel; total length, 23 mm; and needle tip, 15 mm.</td></tr><tr><td>BD OptEIATM&nbsp;TMB Substrate Reagent Set</td><td>BD Biosciences, San Diego, CA, USA</td><td>555214</td><td>We added 100 &mu;L of TMB substrate solution per well.</td></tr><tr><td>Bell Jar</td><td>NA</td><td>NA</td><td>We used a bell jar for anesthesia.</td></tr><tr><td>Clear Flat-Bottom Immuno Nonsterile 96-Well Plates&nbsp;</td><td>ThermoFisher Scientific, Rochester, NY, USA</td><td>442404</td><td>We used Clear Flat-Bottom Immuno Nonsterile 96-Well Plates with MaxiSorp surface treatment for enzyme-linked immunosorbent assays.</td></tr><tr><td>Cotton ball</td><td>Sansho Co., Ltd., Tokyo, Japan</td><td>94-1951</td><td>We used cottons balls to avoid blood contamination. The diameter of cotton balls was 7 mm.</td></tr><tr><td>Fetal bovine serum</td><td>Sigma-Aldrich Co., St. Louis, MO, USA</td><td>172012-500 ML</td><td>We added 10% of fetal bovine serum to the assay diluent.</td></tr><tr><td>Forceps (Micro tweezers for research experiments, curved tip)</td><td>Sansho Co., Ltd., Tokyo, Japan</td><td>91-1506</td><td>We used forceps for disection. Model, HSC 551-10; and overll all length, 105 mm.</td></tr><tr><td>Forceps (Precision tweezers)</td><td>Sansho Co., Ltd., Tokyo, Japan</td><td>91-2772</td><td>We used forceps for disection. Model, HSC 607-12; overll all length, 120 mm; and tip diameter, 0.3 mm.</td></tr><tr><td>Goat Anti-Mouse IgA-UNLB</td><td>SouthernBiotech, Birmingham, AL, USA</td><td>1040-01</td><td>We coated Clear Flat-Bottom Immuno Nonsterile 96-Well Plates with 100 &mu;L/well (20 ng/well) of total immunoglobulin A.</td></tr><tr><td>Isoflurane&nbsp;</td><td>Viatris Inc., Canonsburg, PA, USA</td><td>NA</td><td>We used isoflurane for anesthesia.</td></tr><tr><td>Luminol Rection Reagent Set</td><td>FUJIFILM Wako Pure Chemical Corporation, Osaka, Japan</td><td>124-06511</td><td>We used a luminol reaction reagent set to detect hemoglobin in NLF samples.</td></tr><tr><td>Microtube</td><td>Watson Co., Ltd., Tokyo, Japan</td><td>131-7155C</td><td>We used&nbsp; microtubes for collecting NLF and BALF (1.5-mL microtube with lockining-cap, round bottom, soft touch-cap).</td></tr><tr><td>Milli-Q water</td><td>Merck KGaA, Darmstadt, Germany</td><td>Milli-Q Direct 8</td><td>We used MilliQ water to make a luminol rection solution.</td></tr><tr><td>Mini-Shaker PSU-2T</td><td>Biosan Ltd., Riga, Latvia</td><td>PSU-2T</td><td>We shaked 96-well plates using the Mini-Shaler PSU-2T.</td></tr><tr><td>Mouse strain</td><td>CLEA Japan, Inc., Tokyo, Japan</td><td>NA</td><td>we used 4-month-old C57BL/6 male mice.</td></tr><tr><td>Nunc MicroWell 96-Well, Nunclon Delta-Treated, Flat-Bottom Microplate</td><td>Thermo Fisher Scientific Nunc A/S . Kamstrupvej,Roskilde, Denmark</td><td>136101</td><td>We used the NuncTM MicroWellTM 96-Well, Nunclon Delta-Treated, Flat-Bottom Microplate for the luminol test.</td></tr><tr><td>Peroxidase-conjugated AffiniPure F(ab')2 Fragment Goat Anti-Mouse IgG, F(ab')2 Fragment Specific (minimal cross-reaction to Human, Bovine, and Horse Serum Proteins)</td><td>Jackson ImmunoResearch Laboratories, Inc., West Grove, PA, USA</td><td>115-036-072</td><td>We added 100 &mu;L of peroxidase-conjugated AffiniPure F(ab')2 fragment goat anti-mouse IgG, F(ab')2 fragment specific, as the detection antibody (10,000-fold dilution) to each well.</td></tr><tr><td>Phosphate-buffered saline</td><td>Sigma-Aldrich Co., St. Louis, MO, USA</td><td>D8537-500ML</td><td>We used Dulbecco's Phosphate-Buffered Saline (modified without calcium chloride and magnesium chloride, liquid, sterile-filtered, suitable for cell cullture) for NLF and BALF collection.</td></tr><tr><td>Pipette&nbsp;</td><td>Watson Co., Ltd., Tokyo, Japan</td><td>NA</td><td>We used a 200-&mu;L pipette for nasal lavage.</td></tr><tr><td>Pipette tip</td><td>Watson Co., Ltd., Tokyo, Japan</td><td>NA</td><td>We used a 200-&mu;L multifit pipette tip for nasal lavage.</td></tr><tr><td>Polyoxyethylene(20) Sorbitan Monolaurate (Tween 20)</td><td>FUJIFILM Wako Pure Chemical Corporation, Osaka, Japan</td><td>167-11515</td><td>We added 0.05% of Tween 20 to the washing buffer and 0.2% of Tween 20 to the assay diluent.</td></tr><tr><td>Purified Goat anti-mouse IgG (minimal x-reactivity) Antibody</td><td>BioLegend, San Diego, CA, USA</td><td>405301</td><td>We coated Clear Flat-Bottom Immuno Nonsterile 96-Well Plates with 100 &mu;L/well (10 ng/well) of total immunoglobulin G.</td></tr><tr><td>Small scissors, double-pointed</td><td>Sansho Co., Ltd., Tokyo, Japan</td><td>91-7005</td><td>We used scissors for disection. Model, S-4A; and overll all length, 115 mm.</td></tr><tr><td>Sulfuric acid</td><td>Wako Pure Chemical Industries, Ltd, Tokyo, Japan</td><td>192-04755</td><td>We added 50 &mu;L of 2 N sulfuric acid to each well to stop the reaction.</td></tr><tr><td>Surflo intravenous catheter&nbsp;</td><td>Terumo Corporation, Tokyo, Japan&nbsp;</td><td>SR-FS2032</td><td>We used a surflo intravenous catheter and injected PBS into the trachea for bronchoalveolar lavage. Catheter, 20G; catheter length, 32 mm; catheter inner diameter, 0.80 mm; catheter outer diameter, 1.1 mm; and inner needel, 22G.</td></tr><tr><td>Surgical straight scissors</td><td>Sansho Co., Ltd., Tokyo, Japan</td><td>91-7004</td><td>We used scissors for disection. Model, S-3B; and overll all length, 120 mm.</td></tr><tr><td>Surgilcal plate</td><td>Natsume Seisakusho Co., Ltd., Tokyo, Japan</td><td>KN-321-B</td><td>We used a cork board for dissection. Depth, 20 cm; width, 15 cm; and height 3 cm.</td></tr><tr><td>Synergy H1 Hybrid Multi-Mode Microplate Reader&nbsp;</td><td>Agilent Technologies, Inc., Santa Clara, CA, USA</td><td>NA</td><td>We measured the absorbance of samples at 450 nm, using the Synergy H1 Hybrid Multi-Mode Microplate Reader.</td></tr><tr><td>Syringe</td><td>Terumo Corporation, Tokyo, Japan&nbsp;</td><td>SS-01T</td><td>We used a 1-mL syringe for blood collection from the heart.</td></tr><tr><td>Thread</td><td>&nbsp;Niccho Kogyo Co., Ltd. Tokyo Japan</td><td>NA</td><td>We tied a catheter with thread to ensure proper positioning during the procedure.</td></tr></tbody></table>

verzameling van neusspoelingsvloeistof bij muizen zonder bloedverontreiniging

Murine Nasal Lavage Fluid Collection without Blood Contamination

Verzameling van neusspoelingsvloeistof bij muizen zonder bloedverontreiniging

Upper airway samples, including nasal lavage fluid (NLF) and nasal swabs, are useful in detecting pathogens, including viruses and bacteria, which can ...

murine-nasal-lavage-fluid-collection-without-blood-contamination

Research

JoVE Journal

Immunology and Infection

Immunologie en Infectie

Source: Venuprasad, K., et al. Intra-tracheal Administration of Haemophilus influenzae in Mouse Models to Study Airway Inflammation. J. Vis. Exp. (2016)This video demonstrates the collection and processing of bronchoalveolar lavage fluid from a euthanized mouse infected with non-typeable Haemophilus influenzae for downstream analysis.

Bronchoalveolar Lavage Fluid Collection from a Mouse Model Infected with Non-typeable Haemophilus influenzae

This video demonstrates the use of flow cytometry to identify and count various immune cells in bronchoalveolar lavage fluid. Initially, single immune cells are identified using a viability dye, followed by differential staining of surface marker proteins, enabling the precise enumeration of each cell type.

Identification and Enumeration of Immune Cells in Bronchoalveolar Lavage Fluid by Flow Cytometry

Kolonisatie van de muriene nasopharynx met Streptococcus pneumoniae en de daaropvolgende extractie van aanhangende of gerekruteerde cellen wordt beschreven. Deze techniek omvat het spoelen van de nasopharynx en het opvangen van de vloeistof door de nares en is aanpasbaar voor verschillende uitlezingen, waaronder differentiële celkwantificering en analyse van mRNA-expressie in situ.

Karakterisering van inflammatoire reacties tijdens intranasale kolonisatie met Streptococcus pneumoniae

De gezondheidsstatus van de long wordt weerspiegeld door het type en het aantal immuuncellen die aanwezig zijn in de bronchiolen van de long. We beschrijven een bronchoalveolaire lavage techniek die de isolatie en studie van niet-adherente cellen en oplosbare factoren uit het onderste luchtwegen van muizen mogelijk maakt.

Bronchoalveolaire lavage van meneuze longen om ontstekingscelinfiltratie te analyseren

Dit protocol beschrijft een noninvasive techniek voor de bemonstering van ongestoord mucosal voering vloeistof uit de bovenste luchtwegen. Het kan worden gebruikt voor het uitvoeren van de kwantificering van in vivo niveaus van proteïne bemiddelaars, zoals cytokines en chemokines, bij patiënten van alle leeftijden.

Noninvasive bemonstering van Mucosal voering vloeistof voor de kwantificering van In Vivo bovenste luchtwegen immuun-bemiddelaar niveaus

Dit manuscript beschrijft het gebruik van nasale en bronchiale absorptie technieken, specifiek met behulp van synthetische absorberend matrices (SAM) om te proeven van de mucosal voering vloeistof (MLF) van de bovenste en onderste luchtwegen. Deze methoden bieden betere standaardisatie en verdraagbaarheid dan bestaande respiratoire bemonsteringstechnieken.

Absorptie van vloeistoffen nasale en bronchiale: Precision bemonstering van de menselijke respiratoire Mucosa en laboratorium verwerking van de monsters

Dit protocol beschrijft een verbeterde techniek voor de overvloedige hoeveelheid cerebrospinale vloeistof (CSF) met geen besmetting uit bloed. Met grotere sample collectie en zuiverheid, kunnen meer analyses worden uitgevoerd met behulp van CSF om ons begrip van ziekten die de hersenen en het ruggenmerg beïnvloeden.

Een verbeterde methode voor de verzameling van de cerebrospinale vloeistof van narcose muizen

Deze mededeling beschrijft methodologieën voor isolatie en cultuur van de alveolaire macrofagen van mens en lymfkliertest modellen voor experimentele doeleinden.

Isolatie en In Vitro cultuur van lymfkliertest en menselijke alveolaire macrofagen

Zoutoplossing neusspoeling kan worden gebruikt om de nasale immuunmicro-omgeving van honden te bemonsteren. Omdat de aanpak relatief niet-invasief is en het neusweefsel niet verstoort, kan deze serieel worden uitgevoerd. Cellen en eiwitten die zijn verzameld uit de neusspoeltechniek kunnen worden verwerkt voor verschillende laboratoriumanalyses.

Zoutoplossing voor bemonstering van de nasale immuunmicro-omgeving van honden

Hier presenteren we een protocol voor een geoptimaliseerde bloedafnametechniek bij muizen door middel van venapunctie in het gezicht met behulp van een lancet. We gebruiken deze techniek naast flowcytometrische beoordeling om cellulaire fenotypes van transgene, congenische en reporterstammen te karakteriseren. Het is ook geschikt voor serumcytokine-analyse en diabetesstudies.

Venapunctie in het gezicht voor het afnemen van bloed bij muizen

Leer een methode voor het verzamelen van neusspoelingsvloeistof bij muizen zonder bloedverontreiniging. Deze techniek verbetert de nauwkeurigheid van immunologische analyses en detectie van luchtweginfecties.

Verzameling van neusspoelingsvloeistof bij muizen zonder bloedverontreiniging

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