logo
AAT Bioquest

Protonex™ Red 600-Latex Bead Conjugate

Protonex™ Red-latex bead conjugate demonstrated pH-dependent fluorescence. Unlike most of the existing fluorescent dyes that are more fluorescent at higher pH, acidic conditions enhance the fluorescence of Protonex™ Red-latex bead conjugate. The fluorescence of Protonex™ Red-latex bead conjugate dramatically increases as pH decreases from neutral to the acidic, making it a robust tool to study phagocytosis and its regulation by drugs and/or environmental factors. The lack of fluorescence outside the cell eliminates the wash steps. Protonex™ Red-latex bead conjugate provides a powerful tool to study phagocytosis. Protonex™ Red-latex bead conjugate is low fluorescent outside the cells, but fluoresce brightly red in acidic compartments (such as phagosomes, lysosomes and endosomes). This Protonex™ Red-latex bead conjugate can be also used for multiplexing cell functional analysis with green dyes such as GFP, Fluo-8, calcein, or FITC-labeled antibodies. Protonex™ Red has the spectral properties similar to those of Texas Red, making the common filter set of Texas Red readily available to the assays of Protonex™ Red.

Example protocol

AT A GLANCE

Chemical and Physical Properties

Solvent:

Water

Solids Content:

1% in PBS

Number of Microspheres per mL:

~4e+10

Ex/Em:

575/597 nm

Mean Diameter:

0.72 µm

SAMPLE EXPERIMENTAL PROTOCOL

Important

The following is a recommended protocol for granulocytes. This protocol only provides a guideline and should be modified
according to your specific experimental conditions. 

Protocol
  1. Prepare cells as desired. For example, prepare the granulocytes at 107 cells/mL with Hanks and 20 mM Hepes buffer (HHBS), and add 100 μL to a polypropylene tube.

    Note: Each cell line should be evaluated on an individual basis to determine the optimal cell density. 

  2. Add 1-10 μL of the Protonex™ Red 600-Latex Bead Conjugate to the tube and incubate with gentle shaking for 30 minutes at 37˚C.

    Note: Each cell line should be evaluated on an individual basis to determine the optimal incubation time. 

  3. Prepare an identical sample that is incubated at 4˚C and label it as a control.

  4. At the end of the 30-minute incubation, stop phagocytosis by adding 2mL of ice-cold HHBS and mix well.

  5. Wash the cells 2 times with cold HBSS. 

  6. Resuspend the cells in 500 μL of cold HBSS, keep the samples at 4˚C, and analyze immediately using a fluorescence microscope equipped with a Texas Red® filter set.

    Note: For fluorescence microplate readers, monitor the fluorescence intensity at Ex/Em = 570/600 nm (Cutoff = 585 nm).

Spectrum

Product family

NameExcitation (nm)Emission (nm)
Protonex™ Red 670-Latex Bead Conjugate643660

Citations

View all 3 citations: Citation Explorer
Regulation of IFN-Is by MEF2D Promotes Inflammatory Homeostasis in Microglia
Authors: Lu, Fangfang and Wang, Ronglin and Nie, Tiejian and Gao, Fei and Yang, Shaosong and Huang, Lu and Xia, Li and Shao, Kaifeng and Liu, Jiankang and others,
Journal: (2021)
PHD2 is a regulator for glycolytic reprogramming in macrophages
Authors: Guentsch, Annemarie and Beneke, Angelika and Swain, Lija and Farhat, Katja and Nagarajan, Shunmugam and Wielockx, Ben and Raithatha, Kaamini and Dudek, Jan and Rehling, Peter and Zieseniss, Anke and others,
Journal: Molecular and cellular biology (2017): e00236--16
PHD2 is a regulator for glycolytic reprogramming in macrophages
Authors: Guentsch, Annemarie and Beneke, Angelika and Swain, Lija and Farhat, Katja and Nagarajan, Shunmugam and Wielockx, Ben and Raithatha, Kaamini and Dudek, Jan and Rehling, Peter and Zieseniss, Anke and others, undefined
Journal: Molecular and Cellular Biology (2016): MCB--00236

References

View all 56 references: Citation Explorer
Monitoring phospholipid dynamics during phagocytosis: application of genetically-encoded fluorescent probes
Authors: Sarantis H, Grinstein S.
Journal: Methods Cell Biol (2012): 429
Phagocytosis and digestion of pH-sensitive fluorescent dye (Eos-FP) transfected E. coli in whole blood assays from patients with severe sepsis and septic shock
Authors: Schreiner L, Huber-Lang M, Weiss ME, Hohmann H, Schmolz M, Schneider EM.
Journal: J Cell Commun Signal (2011): 135
The application of fluorescent probes for the analysis of lipid dynamics during phagocytosis
Authors: Flannagan RS, Grinstein S.
Journal: Methods Mol Biol (2010): 121
Quantification of microsized fluorescent particles phagocytosis to a better knowledge of toxicity mechanisms
Authors: Leclerc L, Boudard D, Pourchez J, Forest V, Sabido O, Bin V, Palle S, Grosseau P, Bernache D, Cottier M.
Journal: Inhal Toxicol (2010): 1091
Analysis of macrophage phagocytosis: quantitative assays of phagosome formation and maturation using high-throughput fluorescence microscopy
Authors: Steinberg BE, Grinstein S.
Journal: Methods Mol Biol (2009): 45
Page updated on December 4, 2024

Ordering information

Price
Unit size
Catalog Number21209
Quantity
Add to cart

Additional ordering information

Telephone1-800-990-8053
Fax1-800-609-2943
Emailsales@aatbio.com
InternationalSee distributors
Bulk requestInquire
Custom sizeInquire
Technical SupportContact us
Purchase orderSend to sales@aatbio.com
ShippingStandard overnight for United States, inquire for international
Request quotation

Spectral properties

Excitation (nm)

576

Emission (nm)

597

Storage, safety and handling

H-phraseH303, H313, H333
Hazard symbolXN
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R22

Storage

Refrigerated (2-8 °C); Minimize light exposure
UNSPSC12352200

Platform

Fluorescence microscope

ExcitationTexas Red filter set
EmissionTexas Red filter set
Recommended plateBlack wall, clear bottom
Phagocytosis was examined in RAW 264.7 cells by Protonex™ Red 600-Latex Bead Conjugate (Cat # 21209). The cells were incubated with Protonex™ 600 Latex Beads in growth medium for 4 hours.  CytoTrace™ Green CMFDA  (Cat # 22017) was used to track live cells. The image (20X) was taken using Keyence Fluorescence Microscopy. 
Phagocytosis was examined in RAW 264.7 cells by Protonex™ Red 600-Latex Bead Conjugate (Cat # 21209). The cells were incubated with Protonex™ 600 Latex Beads in growth medium for 4 hours.  CytoTrace™ Green CMFDA  (Cat # 22017) was used to track live cells. The image (20X) was taken using Keyence Fluorescence Microscopy. 
Phagocytosis was examined in RAW 264.7 cells by Protonex™ Red 600-Latex Bead Conjugate (Cat # 21209). The cells were incubated with Protonex™ 600 Latex Beads in growth medium for 4 hours.  CytoTrace™ Green CMFDA  (Cat # 22017) was used to track live cells. The image (20X) was taken using Keyence Fluorescence Microscopy.