Flow cytometry Anti-bodies Mouse - CD206

Flow cytometry is an immunophenotyping technique whereby sing cell suspensions are stained for either cell surface markers or intracellular proteins by fluorescently-labelled antibodies and analyzed with a flow cytometer, where fluorescently-labelled molecules are excited by the laser to emit light at varying wavelengths, which is then detected by the instrument. There are several key criteria which are required to be kept in mind while designing a flow experiment- 1. Antibody titration (optimal dilution of antibodies should be calculated in order to avoid over- or under- saturated signals for proper detection of surface and intracellular markers), 2. Precision (3 or more replicates of the sample should be used per experiment), 3. Specificity (proper isotype controls should be included in the experiment), 4. Day-to-day variability (experiments should be repeated 3 or more times to ensure consistency and avoid variability due to flow cytometer settings), 5. Antibody interaction (Fluorescence minus one or FMO should be used, which is the comparison of signals from panel minus one antibody vs. the full panel), and 6. Antibody stability (fluorescently-labelled antibodies should be stored at 4C).

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Found 3 matching solutions for this experiment

Alexa Fluor® 647 Rat Anti-Mouse CD206

BD Biosciences

Upstream tips	Protocol tips	Downstream tips
Mouse lung alveolar macrophages were collected by fitting a venous indwelling needle into a mouse trachea. PBS was injected to wash the lung six consecutive times (0.8 ml/wash). Total bronchial alveolar lavages were collected and washed with PBS.	In brief, cells were washed once in room temperature flow buffer (PBS supplemented with 1% (v/v) FBS and 0.05% NaN3) and once in ice‐cold flow buffer, and cell staining was performed on ice. Samples were analysed using a BD Cyan flow cytometer using CellQuest software (BD Biosciences, San Jose, CA, USA).	). Further analysis was performed using FloJo software (Tree Star Inc., Ashland, OR, USA).

Upstream tips
Mouse lung alveolar macrophages were collected by fitting a venous indwelling needle into a mouse trachea. PBS was injected to wash the lung six consecutive times (0.8 ml/wash). Total bronchial alveolar lavages were collected and washed with PBS.
Protocol tips
In brief, cells were washed once in room temperature flow buffer (PBS supplemented with 1% (v/v) FBS and 0.05% NaN3) and once in ice‐cold flow buffer, and cell staining was performed on ice. Samples were analysed using a BD Cyan flow cytometer using CellQuest software (BD Biosciences, San Jose, CA, USA).
Downstream tips
). Further analysis was performed using FloJo software (Tree Star Inc., Ashland, OR, USA).

Manufacturer protocol Publication protocol 1 Relevant paper

APC anti-mouse CD206 (MMR) Antibody

BioLegend

Upstream tips	Protocol tips	Downstream tips
Single-cell suspensions were incubated with a Fc receptor block (1 μg/1 × 106 cells; BD Bioscience, San Diego, CA) to reduce nonspecific anti-body binding.	Dead cells were excluded using a Live/Dead Fixable Blue Dead Cell Stain Kit (Invitrogen) or 4,6-diamidino-2-phenylindole dihydrochloride (DAPI) (Sigma-Aldrich, St. Louis, MO).	Flow cytometry was performed using BD LSR II and BD FACS Aria III flow cytometers (BD Bioscience), and data were analyzed with FlowJo software (TreeStar, Ashland, OR).

Upstream tips
Single-cell suspensions were incubated with a Fc receptor block (1 μg/1 × 106 cells; BD Bioscience, San Diego, CA) to reduce nonspecific anti-body binding.
Protocol tips
Dead cells were excluded using a Live/Dead Fixable Blue Dead Cell Stain Kit (Invitrogen) or 4,6-diamidino-2-phenylindole dihydrochloride (DAPI) (Sigma-Aldrich, St. Louis, MO).
Downstream tips
Flow cytometry was performed using BD LSR II and BD FACS Aria III flow cytometers (BD Bioscience), and data were analyzed with FlowJo software (TreeStar, Ashland, OR).

Manufacturer protocol Publication protocol 1 Relevant paper

CD206 antibody | MR5D3

Bio-Rad Laboratories

Upstream tips	Protocol tips	Downstream tips
	An isotype control antibody was used as a negative control.	The experiments were performed using a FACS Diva Version 6.1.2 automated cell analyzer (BD FACS Canto II).

Protocol tips
An isotype control antibody was used as a negative control.
Downstream tips
The experiments were performed using a FACS Diva Version 6.1.2 automated cell analyzer (BD FACS Canto II).

Manufacturer protocol Publication protocol 1 Relevant paper

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