小鼠支气管肺泡灌洗

Fan  Sun; Gutian  Xiao; Zhaoxia  Qu

doi:10.21769/BioProtoc.2287

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Peer-reviewed

Murine Bronchoalveolar Lavage

小鼠支气管肺泡灌洗

FS Fan Sun

GX Gutian Xiao

ZQ Zhaoxia Qu email

发布: 2017年05月20日第7卷第10期 DOI: 10.21769/BioProtoc.2287 浏览次数: 25765

评审: HongLok LungRiddhi Atul JaniAnonymous reviewer(s)

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参见作者原研究论文

The authors used this protocol in:

Cover of Oncogene, featuring study using the protocol.

May 2016

实验方案合集

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Abstract

A basic Bronchoalveolar lavage (BAL) procedure in mouse is described here. Cells and fluids obtained from BAL can be analyzed by Hema3-staining, immunostaining, Fluorescence-activated cell sorting (FACS), PCR, bicinchoninic acid protein assay, enzyme-linked immunosorbent assay (ELISA), luminex assays, etc., to examine the immune cells, pathogens, proteins such as cytokines/chemokines, and the expression levels of inflammation-related and other genes in the cells. This will help to understand the underlying mechanisms of these lung diseases and develop specific and effective drugs.

Keywords: Bronchoalveolar lavage (支气管肺泡灌洗)

Lung cancer (肺癌)

Hema3-staining (Hema3染色)

Background

Bronchoalveolar lavage (BAL) is a simple and typical method commonly performed to diagnose pulmonary diseases including lung cancer (Daubeuf and Frossard, 2012). It is used to sample pulmonary components to determine the protein composition, immune cells and pathogens in the lung. Pulmonary chronic inflammation plays a critical role in lung cancer initiation and progression. To clarify the underlying mechanism of inflammation in lung tumorigenesis, a basic BAL protocol in mice is used in our laboratory to determine the pulmonary immune response (Qu et al., 2015; Zhou et al., 2015; Sun et al., 2016; Zhou et al., 2017).

Materials and Reagents

Needles (BD, catalog number: 305167 ) or tapes
Nylon string (Dynarex, catalog number: 3243 )
22 G x 1” Exel Safelet Catheter (Exel International, catalog number: 26746 )
1 ml syringe (BD, catalog number: 309659 )
1.5 ml Eppendorf tubes (VWR, catalog number: 87003-294 )
0.22 µm filter (EMD Millipore, catalog number: SLGP033RS )
Mice (THE JACKSON LABORATORY)
70% ethanol (Decon Labs, catalog number: 2701 )
Protease inhibitor cocktail (Roche Diagnostics, catalog number: 11697498001 )
Phenylmethylsulfonyl fluoride (PMSF) (Thermo Fisher Scientific, Thermo Scientific^TM, catalog number: 36978 )
Sodium chloride (NaCl) (Sigma-Aldrich, catalog number: S9625 )
Potassium chloride (KCl) (Sigma-Aldrich, catalog number: P9541 )
Disodium hydrogen phosphate heptahydrate (Na₂HPO₄·7H₂O) (Fisher Scientific, catalog number: BP331-500 )
Potassium phosphate monobasic (KH₂PO₄) (Acros Organics, catalog number: 205925000 )
Ammonium chloride (NH₄Cl) (Sigma-Aldrich, catalog number: A9434-1KG )
Potassium bicarbonate (KHCO₃) (Sigma-Aldrich, catalog number: P9144-1KG )
Note: This product has been discontinued.
Ethylenediaminetetraacetic acid disodium salt dihydrate (Na₂-EDTA-2H₂O) (Sigma-Aldrich, catalog number: E5134-100G )
Phosphate-buffered saline (PBS) (see Recipes)
ACK lysis buffer (see Recipes)