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Phagocytosis Detection Kit Red

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Regulatory Status
RUO
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a. Phagocytosis-Detection-Kit_Live-Cell-Imaging-_082729.
Live-cell fluorescence imaging of RAW264.7 cells treated with with Phagocytosis Detection Beads (red) in complete growth media for 4 hours. Cells were then stained with Viability Dye Green (green) for 30 minutes and nuclei were stained with Hoechst 33342 (blue). Images were captured using a 60X objective. Scale bar: 10 µm
  • a. Phagocytosis-Detection-Kit_Live-Cell-Imaging-_082729.
    Live-cell fluorescence imaging of RAW264.7 cells treated with with Phagocytosis Detection Beads (red) in complete growth media for 4 hours. Cells were then stained with Viability Dye Green (green) for 30 minutes and nuclei were stained with Hoechst 33342 (blue). Images were captured using a 60X objective. Scale bar: 10 µm
  • b. Phagocytosis-Detection-Kit_FC-_082729.
    THP-1 cells were treated with Phagocytosis Detection Beads and Viability Dye Green for 1 hour (filled histogram) or left untreated (open histogram). Cells were analyzed on flow cytometer using PE channel.
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421938 1 kit £278
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Description

Phagocytosis is a complex process for the ingestion and elimination of pathogens. It is also important for the elimination of apoptotic cells, hence fundamental for tissue homeostasis. Phagocytosis consists of recognition and ingestion of particles larger than 0.5 µm into a plasma membrane-derived vesicle, known as phagosome. Professional phagocytes include monocytes, macrophages, neutrophils, dendritic cells, osteoclasts, and eosinophils. These cells are in charge of eliminating microorganisms and presenting them to cells of the adaptive immune system. In addition, fibroblasts, epithelial cells, and endothelial cells can also perform phagocytosis. These nonprofessional phagocytes cannot ingest microorganisms but are important in eliminating apoptotic bodies. Once internalized, the phagosome fuse with lysosomes to form secondary phagolysosomes for digestion, resulting in progressive decrease of pH. The Phagocytosis Detection Beads are non-fluorescent
outside of cells. However, its fluorescence dramatically increases as they are internalized and within the acidic phagosomes/phagolysosome compartments.

Product Details
Technical Data Sheet (pdf)

Product Details

Verified Reactivity
Human, Mouse
Reported Reactivity
All Species
Formulation
1 vial Phagocytosis Detection Beads, 15 µL
1 vial Viability Dye Green, 20 µL
1 vial DMSO, 100 µL
Storage & Handling
4°C
Application

FC - Quality tested
Live cell imaging - Verified
Recommended Usage

Refer to protocol

Application Notes

Component:
1 vial (15 µL) Phagocytosis Detection Beads (2-8°C, protected from light)
1 vial (20 µL) Viability Dye Green (2-8°C, protected from light)
1 vial (100 µL) DMSO (room temperature)
Note: all unused stock solutions should be divided into single-use aliquots and stored at -20°C after preparation. Avoid repeated freeze-thaw cycyles.

Required Materials Not Included:
Buffer: Phosphate-buffered saline (PBS), phenol-free media, or HBSS
Complete growth media (cell type dependent)

Detection/Imaging Guidelines:
Phagocytosis Detection Beads
Ex/Em = 570/600 nm
Fluorescence microscope filter set: Texas Red
Flow cytometry channel: PE with yellow/green laser excitation

Viability Dye Green
Ex/Em = 490/525 nm
Fluorescence microscope filter set: FITC
Flow cytometry channel: FITC

Prepare Reagents:

  1. Bring all three components to room temperature: Phagocytosis Detection Beads, Viability Dye Green, and DMSO
    Note: all unused stock solutions should be divided into single-use aliquots and stored at -20°C after preparation. Avoid repeated freeze-thaw cycyles.
  2. Viability Dye Green: Prepare stock solution of Viability Dye Green by adding 20 µL of DMSO.
  3. Then, prepare the desired amount of Viability Dye Green working solution in preferred complete growth media by adding 5 µL of the stock solution to 2 mL growth media. Mix well.
    Note: all unused stock solutions can be aliquoted into single-use vials and stored at -20°C, protexted from light.
  4. Phagocytosis Detection Beads: Prepare desired volume of Phagocytosis Detection Beads working solution in complete growth media by adding 4 µL for every 1 mL growth media. Mix well.

Live-Cell Imaging Assay Protocol:

  1. Add 16 µL of Phagocytosis Detection Beads working solution to every 200 µL of your preferred complete media. You may add the Phagocytosis Detection Beads working solution directly into each well of adherent cells in complete media. Cells should be about 80% confluent.
    Note: Final concentration may need to be optimized by end user for certain cell types and assay conditions. 
  2. Incubate cells at 37°C for 4 hours.
    Note: The incubation time may need to be adjusted according to cell type and assay conditions.
  3. Add 16 µL of the Viability Dye Green working solution to every 200 µL media in cell culture wells.
    Note: Final concentration may need to be optimized by end user for certain cell types and assay conditions.
  4. Optional: include live-cell nuclear counterstain such as DRAQ5™ (Cat. No. 424101).
  5. Incubate cells at 37°C for 30 minutes.
  6. Gently wash cells twice with PBS, phenol-free media or HBSS.
  7. Add sufficient amount of PBS, phenol-free media or HBSS to the wells and proceed immediately to livecell imaging.
Flow Cytometry Assay Protocol:
  1. Resuspend cells (1-2 x 106) in 200 µL of PBS, HBSS, or Cell Staining Buffer (Cat. No. 420201).
  2. Add 16 µL of Phagocytosis Detection Beads working solution to the cell suspension. 
    Note: Final concentration may need to be optimized by end user for certain cell types and assay conditions. You may also prepare single color tubes for cell only, PE, and FITC for compensation.
  3. Add 16 µL of Viability Dye Green working solution to the cell suspension.
    Note: Final concentration may need to be optimized by end user for certain cell types and assay conditions.
  4. Incubate at 37°C for 1 hour.
  5. Wash cells twice with PBS, HBSS or Cell Staining Buffer. After the second wash, gently remove supernatant from the cell pellet.
  6. Resuspend in 200-300 µL of PBS, HBSS, or Cell Staining Buffer
  7. Proceed to flow cytometry analysis using an appropriate filter set.

Antigen Details

Distribution

Phagocytes

Function
Phagocytosis activity
Cell Type
Dendritic cells, Macrophages
Biology Area
Cell Biology, Immunology
Molecular Family
Organelle Markers
Gene ID
NA

Related FAQs

There are no FAQs for this product.
Go To Top Version: 2    Revision Date: 02/24/2025

For Research Use Only. Not for diagnostic or therapeutic use.

 

This product is supplied subject to the terms and conditions, including the limited license, located at www.biolegend.com/terms) ("Terms") and may be used only as provided in the Terms. Without limiting the foregoing, BioLegend products may not be used for any Commercial Purpose as defined in the Terms, resold in any form, used in manufacturing, or reverse engineered, sequenced, or otherwise studied or used to learn its design or composition without express written approval of BioLegend. Regardless of the information given in this document, user is solely responsible for determining any license requirements necessary for user’s intended use and assumes all risk and liability arising from use of the product. BioLegend is not responsible for patent infringement or any other risks or liabilities whatsoever resulting from the use of its products.

 

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Toll-Free Phone: 1-877-Bio-Legend (246-5343) Phone: (858) 768-5800 Fax: (877) 455-9587

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