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[pH probe for Acidic Organelle Imaging] AcidiFluor™ ORANGE

  • High S/N ratio
  • Orange color fluorescence and usable for multicolor imaging
  • Great photostability

AcidiFluor™ ORANGE is a fluorescence imaging probe which enhances fluorescence dramatically in acidic environments. This probe can stain acidic organelles such as lysosome, late endosome and granule selectively. Its excellent selectivity enables to detect acidic environment. Namely, in the condition of pH5.0, identical to the environment of acidic organelles, fluorescence intensity is 50-fold or more compared to physical pH 7.4. AcidiFluor™ ORANGE shows great photostability against irradiating excitation light. As it emits orange color fluorescent by excitation at 532nm or 514 nm, multicolor imaging is possible by combining with blue fluorescence (CFP, Hoechst, etc.), green fluorescence (GFP, fluorescein, etc.) and near-infrared fluorescence. AcidiFluor™ ORANGE can be used for detecting acidic organelles, observing granule release, imaging of endocytosis/exocytosis and so on.

Code No. Product Size Price Protocol MSDS
GC301 AcidiFluor™ ORANGE 10μg×20 $680.00 Donwload
GC3011 AcidiFluor™ ORANGE 10μg×10 $380.00

For more cell applications, see application note

Feature of AcidiFluor™ ORANGE

λabs 535 nm λfl 560 nm

pKa 5.1

1. High S/N ratio

Fluorescent spectra of AcidiFluor ORANGE(TM) dependent on pH
Figure 1. Fluorescence spectra of AcidiFluor Orange as a function of pH.

Fluorescence spectra of AcidiFluor Orange was measured in phosphate buffer pH 5.0 or pH 7.4, respectively. pH 5.0 buffer corresponds to the condition in the acidic organelles, and pH 7.4 corresponds to the physiological cytoplasmic conditions. Fluorescent intensity of AcidiFluor Orange at pH 5.0 increased 50 fold to that in the pH 7.4 buffer. ( λex 532 nm / λem 568 nm )

2. Emits orange color fluorescence, usable for multicolor imaging

Example of multicolor imaging using HeLa cell
Fig.2 Example of multicolor imaging using HeLa cell

HeLa cell expressed Mitochondria-GFP was stained multiplicity with AcidiFluor™ ORANGE and Hoechst33342. Lysosomes were stained orange with AcidiFluor™ ORANGE, nucleuses were stained blue with Hoechst33342 and Mitochondria were stained green with GFP. As shown in Fig.2, AcidiFluor™ ORANGE can be useful for multicolor imaging.

3. Great photostability

Comparison to competitor’s products by photocleaching test

Comparison to competitor’s products by photocleaching test
Fig.3 Comparison to competitor’s products by photocleaching test

Samples were continuously illuminated for 180 seconds and viewed on a confocal microscopy. Although LysoTracker® Green DND-26 and LysoTracker® Red DND-99 discolored dramatically, fluorescence of AcidiFluor™ ORANGE remained. LysoSensor™ Green DND-189 was not suitable for continuous observation because fluorescence leaked to cytoplasm by excitation light irradiation.

Goryo Chemical Company commercialized AcidiFluor™ ORANGE under the guidance of Prof. Kenzo Hirose (Professor of Graduate School of Medicine, The University of Tokyo, Department of Neurobiology). AcidiFluor™ ORANGE was licensed from Tokyo University. The development of this product was supported by JST (Japan Science and Technology Agency) program “Development of Systems and Technology for Advanced Measurement and Analysis”. Please refer to the application note.

Kitakaze K, Mizutani Y, Sugiyama E, Tasaki C, Tsuji D, Maita N, Hirokawa T, Asanuma D, Kamiya M, Sato K, Setou M, Urano Y, Togawa T, Otaka A, Sakuraba H, Itoh K.
Protease-resistant modified human β-hexosaminidase B ameliorates symptoms in GM2 gangliosidosis model 
J Clin Invest. 2016 Mar 28. pii: 85300. doi: 10.1172/JCI85300.
Hayashi A, Asanuma D, Kamiya M, Urano Y, Okabe S. High affinity receptor labeling based on basic leucine zipper domain peptides conjugated with pH-sensitive fluorescent dye: Visualization of AMPA-type glutamate receptor endocytosis in living neurons Neuropharmacology Volume 100, January 2016, 66-75
Masayuki Isa, Daisuke Asanuma, Shigeyuki Namiki, Kazuo Kumagai, Hirotatsu Kojima, Takayoshi Okabe, Tetsuo Nagano, and Kenzo Hirose, ACS Chem Biol 2014 Oct 22;9(10):2237-41, “High-throughput screening system to identify small molecules that induce internalization and degradation of HER2.”
Watanabe R, Soga N, Fujita D, Tabata KV, Yamauchi L, Kim SH, Asanuma D, Kamiya M, Urano Y, Suga H and Noji H. Nature Communications 2014 Jul24; 5, Article number: 4519 doi:10.1038/ncomms5519 “Arrayed lipid bilayer chambers allow single-molecule analysis of membrane transporter activity”
Asanuma D, Takaoka Y, Namiki S, Takikawa K, Kamiya M, Nagano T, Urano Y & Hirose K. Acidic-pH-Activatable Fluorescence Probes for Visualizing Exocytosis Dynamics. Angew Chem Int Ed 2014, doi:10.1002/anie.201402030.

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