| Size | Price | Stock | Qty |
|---|---|---|---|
| 1mg |
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| 5mg | |||
| Other Sizes |
| Targets |
Peroxynitrite (ONOO-) is the primary molecular target. The probe is designed for the selective detection of ONOO- over other reactive oxygen species (ROS) and reactive nitrogen species (RNS).
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|---|---|
| ln Vitro |
1. Preparation of HKGreen-4I working solution 1.1 Preparation of stock solution: Dilute 1 mg of HKGreen-4I with 150 μL of anhydrous DMSO to prepare a 10 mM stock solution. It is advised to aliquot and store HKGreen-4I storage solution at -20°C or -80°C in the dark. 1.2 Creating a functional solution Use preheated serum-free cell culture medium or PBS dilution stock solution to prepare a 1-10 μM HKGreen-4I working solution. Note: Please prepare the HKGreen-4I working fluid for usage by adjusting its concentration to suit the current circumstances. 2. Cell staining (suspension cells) 2.1 Centrifuge the cells, add PBS, and wash twice for five minutes each time. The density of cells is 1×106/mL. 2.2 Add 1 mL of the HKGreen-4I working solution, then let it sit at room temperature for a duration of 5 to 30 minutes. 2.3 Centrifuge for 3–4 minutes at 400 g, then remove the supernatant. 2.4 After adding PBS, wash the cells twice for five minutes each. 2.5 Re-suspend the cells in 1 milliliter of PBS or serum-free media, and use a flow cytometer or fluorescence microscope to observe. 3. Staining adherent cells in cells 3.1 Use sterile coverslips to cultivate adherent cells. 3.2 Aspirate extra culture medium after removing the coverslip from the medium. 3.3 Cover the cells fully with 100 μL of the dye working solution, shake gently, and allow incubate for five to thirty minutes. 3.4 Use a flow cytometer or fluorescence microscope to observe after aspirating the dye working solution and washing it twice in culture media for a total of five minutes each time.
HKGreen-4I exhibits high sensitivity and selectivity for ONOO- in cell-free assays. Upon reaction with ONOO-, the probe undergoes a specific chemical transformation that results in a strong fluorescence turn-on signal. Its excitation and emission maxima are 520 nm and 543 nm, respectively. |
| ln Vivo |
There is no specific in vivo activity data available for this compound. However, as a fluorescent probe for reactive species, similar compounds are typically used for real-time imaging of ONOO- in live cells and animal models.
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| Enzyme Assay |
A standard protocol for testing ONOO- detection involves incubating HKGreen-4I (e.g., 5 uM) with varying concentrations of ONOO- in a buffer solution (e.g., PBS, pH 7.4) for a short period (e.g., 5-10 minutes) at room temperature. The fluorescence intensity (Ex/Em: 520/543 nm) is then measured with a spectrophotometer.
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| Cell Assay |
A general protocol for cellular imaging involves seeding cells (e.g., macrophages) in a confocal dish and incubating with HKGreen-4I (e.g., 1-10 uM) for 20-30 minutes at 37degC. Cells are then washed to remove excess probe. Fluorescence imaging is performed with a confocal microscope using excitation at 520 nm and emission collection at 543 nm.
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| Animal Protocol |
A typical protocol for in vivo imaging involves intravenous or intraperitoneal injection of an optimized concentration of HKGreen-4I into a live animal model. After a suitable circulation time (e.g., 30 minutes to 2 hours), the animal is placed under an in vivo imaging system to monitor ONOO--induced fluorescence.
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| ADME/Pharmacokinetics |
General PK properties for small molecule probes are not applicable, as these compounds are typically used for acute imaging. However, formulation can be in PBS or saline with co-solvents like DMSO for in vivo use.
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| Toxicity/Toxicokinetics |
General toxicity for imaging probes is considered low. For HKGreen-4I, no specific toxicity studies are reported. In similar experiments, incubation with up to 10 uM for 24 hours in cell models shows good biocompatibility and no significant cytotoxicity.
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| References | |
| Additional Infomation |
HKGreen-4I is a research tool for studying oxidative stress and inflammatory diseases. Its ONOO--specific turn-on response offers a low background, making it ideal for high-contrast imaging. This product is for research use only and not for diagnostic or therapeutic applications.
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| Molecular Formula |
C36H30N2O11
|
|---|---|
| Molecular Weight |
666.630210399628
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| Exact Mass |
666.184
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| CAS # |
1448821-82-6
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| PubChem CID |
89717569
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| Appearance |
White to pink solid powder
|
| LogP |
4.5
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| Hydrogen Bond Donor Count |
1
|
| Hydrogen Bond Acceptor Count |
12
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| Rotatable Bond Count |
11
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| Heavy Atom Count |
49
|
| Complexity |
1250
|
| Defined Atom Stereocenter Count |
0
|
| SMILES |
O1C(C2C=C(C(N(CC(=O)OC)CC(=O)OC)=O)C=CC=2C21C1C=CC(=CC=1OC1C=C(C=CC2=1)N(C)C1C=CC(=CC=1)O)OC(C)=O)=O
|
| InChi Key |
ZOZXUNDNNSKANR-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C36H30N2O11/c1-20(39)47-25-11-14-29-31(17-25)48-30-16-23(37(2)22-6-9-24(40)10-7-22)8-13-28(30)36(29)27-12-5-21(15-26(27)35(44)49-36)34(43)38(18-32(41)45-3)19-33(42)46-4/h5-17,40H,18-19H2,1-4H3
|
| Chemical Name |
methyl 2-[[3'-acetyloxy-6'-(4-hydroxy-N-methylanilino)-3-oxospiro[2-benzofuran-1,9'-xanthene]-5-carbonyl]-(2-methoxy-2-oxoethyl)amino]acetate
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| HS Tariff Code |
2934.99.9001
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| 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)
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| Solubility (In Vitro) |
DMSO: 40 mg/mL (60.00 mM)
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|---|---|
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 6 mg/mL (9.00 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 60.0 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: 3 mg/mL (4.50 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 30.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly. Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution. View More
Solubility in Formulation 3: ≥ 1.75 mg/mL (2.63 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution. |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 1.5001 mL | 7.5004 mL | 15.0008 mL | |
| 5 mM | 0.3000 mL | 1.5001 mL | 3.0002 mL | |
| 10 mM | 0.1500 mL | 0.7500 mL | 1.5001 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.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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