ATP-Red 1 is a multi-site binding switchable fluorescent probe that can selectively and quickly detect ATP content in living cells.

Physicochemical Properties

Molecular Formula	C34H36BN3O4
Molecular Weight	561.478348731995
Exact Mass	561.279
CAS #	1847485-97-5
PubChem CID	132274056
Appearance	Pale purple to purple solid powder
Hydrogen Bond Donor Count	2
Hydrogen Bond Acceptor Count	6
Rotatable Bond Count	8
Heavy Atom Count	42
Complexity	893
Defined Atom Stereocenter Count	0
SMILES	O1C2C=C(C=CC=2C2(C3C=CC(=CC1=3)N(CC)CC)C1C=CC=CC=1C(N2C1C=CC=CC=1B(O)O)=O)N(CC)CC
InChi Key	SSNKNUIFMZUJTL-UHFFFAOYSA-N
InChi Code	InChI=1S/C34H36BN3O4/c1-5-36(6-2)23-17-19-27-31(21-23)42-32-22-24(37(7-3)8-4)18-20-28(32)34(27)26-14-10-9-13-25(26)33(39)38(34)30-16-12-11-15-29(30)35(40)41/h9-22,40-41H,5-8H2,1-4H3
Chemical Name	[2-[3',6'-bis(diethylamino)-3-oxospiro[isoindole-1,9'-xanthene]-2-yl]phenyl]boronic acid
Synonyms	ATP-Red 1; 1847485-97-5; [2-[3',6'-bis(diethylamino)-3-oxospiro[isoindole-1,9'-xanthene]-2-yl]phenyl]boronic acid; (2-(3',6'-Bis(diethylamino)-3-oxospiro[isoindoline-1,9'-xanthen]-2-yl)phenyl)boronic acid;
HS Tariff Code	2934.99.9001
Storage	Powder-20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month Note: This product requires protection from light (avoid light exposure) during transportation and storage.
Shipping Condition	Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Biological Activity

Targets	Fluorescent probe/dye
ln Vitro	ATP-Red 1 is a switchable fluorescent probe with multiple binding sites that may respond selectively and quickly to ATP levels in living cells. The highest absorption and emission wavelengths are 570/566 and 590/585 nm, respectively. ATP-Red 1 has strong membrane permeability, and its fluorescence intensity increases by 5.6 times in the presence of 5 mM ATP. After KCN inhibited OXPHOS, ATP-Red 1 (2.5 μM, 20 minutes) showed lower fluorescence, resulting in reduced mitochondrial ATP levels in OSCC cells [1].
Cell Assay	1. Solution Preparation 1.1 Stock Solution Preparation • Solvent: DMSO • Concentration: Recommended 10 mM • Storage: Aliquot and store at -20°C or -80°C, protected from light. Avoid repeated freeze-thaw cycles. 1.2 Working Solution Preparation • Dilute the stock solution in PBS or serum-free cell culture medium to a final concentration of 1–10 µM (optimize based on experimental conditions). • Note: Prepare working solution immediately before use and protect from light. ________________________________________ 2. Cell Staining Protocol 2.1 Cell Preparation • Suspend cells in 100 µL PBS in a 96-well plate at a density of 2×10⁵ cells per well. 2.2 ATP-Red 1 Staining • Add ATP-Red 1 to achieve a final concentration of 5–10 µM. 2.3 Incubation • Incubate cells at room temperature or 37°C for 15 min (extend to 30 min for live-cell imaging). 2.4 Solution Removal • Remove the ATP-Red 1 working solution after incubation. 2.5 Fluorescence Detection • Analyze using a fluorescence microscope (Ex/Em = 510/590 nm).
References	[1]. A Multisite-Binding Switchable Fluorescent Probe for Monitoring Mitochondrial ATP Level Fluctuation in Live Cells. Angew Chem Int Ed Engl. 2016 Jan 26;55(5):1773-6.
Additional Infomation	Adenosine triphosphate (ATP), commonly produced in mitochondria, is required by almost all the living organisms; thus fluorescent probes for monitoring mitochondrial ATP levels fluctuation are essential and highly desired. Herein, we report a multisite-binding switchable fluorescent probe, ATP-Red 1, which selectively and rapidly responds to intracellular concentrations of ATP. Live-cell imaging indicated that ATP-Red 1 mainly localized to mitochondria with good biocompatibility and membrane penetration. In particular, with the help of ATP-Red 1, we successfully observed not only the decreased mitochondrial ATP levels in the presence of KCN and starvation state, but also the increased mitochondrial ATP levels in the early stage of cell apoptosis. These results indicate that ATP-Red 1 is a useful tool for investigating ATP-relevant biological processes. [1]

Solubility Data

Solubility (In Vitro)

DMSO : ~50 mg/mL (~89.05 mM)

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 2.5 mg/mL (4.45 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. Solubility in Formulation 2: ≥ 2.5 mg/mL (4.45 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	1.7810 mL	8.9050 mL	17.8101 mL
	5 mM	0.3562 mL	1.7810 mL	3.5620 mL
	10 mM	0.1781 mL	0.8905 mL	1.7810 mL

*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.