| Size | Price | Stock | Qty |
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| 5mg |
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| 10mg |
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| Other Sizes |
| Targets |
ELF97 phosphate is a substrate for enzymes in the phosphatase family, including alkaline phosphatase (ALP) and acid phosphatase. It does not have a therapeutic target. Its molecular target is the active site of the phosphatase enzyme. The enzyme's nucleophilic cysteine residue attacks the phosphorus atom, cleaving the phosphate group and generating the fluorescent precipitate. This specificity makes it an ideal tool for detecting enzyme activity in situ.
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| ln Vitro |
In vitro, the kinetic parameters of ELF97 phosphate have been characterized using purified alkaline phosphatase. The substrate hydrolysis follows Michaelis-Menten kinetics with a Km of approximately 2-5 uM. It is not an inhibitor; it is a reporter substrate. The EC₅0 is not applicable. The fluorescence signal (Ex/Em: 360/530 nm) is stable for hours, enabling quantitative assays. The substrate itself is non-cytotoxic and non-toxic at working concentrations (1-100 uM).
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| ln Vivo |
In vivo, ELF97 phosphate is not a therapeutic agent. It is an in vivo research tool for detecting phosphatase activity in live tissues. It can be applied topically or injected locally (e.g., intradermally, intraperitoneally) in animal models. In living mice, it is used to visualize sites of bone formation (where ALP is active) or tumors. However, it does not treat any disease. Its primary in vivo use is as a histochemical stain for whole-mount or sectioned tissues.
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| Enzyme Assay |
No cell-free binding assays are performed. Cell-free enzyme kinetic assays are used. For ALP assay, 10 uL of ELF97 phosphate working solution (100 uM) is mixed with 100 uL of purified ALP (1-10 U/L) in 0.1 M Tris-HCl (pH 9.0) containing 1 mM MgCl2. The reaction is incubated at 37degC for 10-30 min. The reaction is stopped by adding 10 uL of 0.5 M EDTA (pH 8.0). Fluorescence is measured in a plate reader (excitation 360 nm, emission 530 nm). A standard curve is prepared using known concentrations of ELF97 alcohol.
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| Cell Assay |
For cell-based assays, cells are cultured in 96-well plates or on coverslips. For detection of endogenous phosphatase, cells are fixed (e.g., with 4% paraformaldehyde) or used live. Cells are washed with PBS and incubated with ELF97 phosphate (10-100 uM) in appropriate buffer (e.g., 0.1 M Tris-HCl, pH 9.0 for ALP, or 0.1 M acetate buffer, pH 5.0 for acid phosphatase) for 30-60 min at 37degC. After incubation, cells are washed and the yellow-green precipitate is imaged by fluorescence microscopy (using a Texas Red or FITC filter). It is not used for viability assays.
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| Animal Protocol |
No animal efficacy studies are performed. For in vivo imaging, a mouse model is injected intraperitoneally or intravenously with the substrate, or the substrate is applied topically. For example, in a mouse model of heterotopic ossification, the animal is euthanized, and the tissue is excised and incubated in ELF97 phosphate solution. Alternatively, in live zebrafish embryos, the substrate can be added to the water to detect notochord ALP activity. The outcome is qualitative or semi-quantitative imaging data.
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| ADME/Pharmacokinetics |
No traditional PK data (Cmax, Tmax, AUC) are collected for this compound because it is not a drug. It is a histochemical substrate that is typically used in fixed or euthanized tissues. If administered systemically, it would be rapidly cleared or hydrolyzed by plasma phosphatases. In zebrafish embryos (0.1-1 uM), it is stable for 1-2 h before photobleaching occurs. For fixed tissue assays, the reaction is stopped to stabilize the signal. It is generally considered to have negligible bioavailability for cellular entry.
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| Toxicity/Toxicokinetics |
ELF97 phosphate is considered non-toxic and non-hazardous at recommended working concentrations (1-100 uM). The acute toxicity (LD₅0) has not been determined, as it is not administered systemically. In cell viability assays (MTT or Alamar Blue) using HeLa or HEK293 cells, concentrations up to 500 uM for 24 h show no significant reduction in viability. It is not a skin or eye irritant. It is not classified as a hazardous substance (GHS). Standard laboratory precautions (gloves, lab coat) are adequate.
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| References | |
| Additional Infomation |
Elf 97 is an aryl phosphate ester, belonging to the quinazoline and monochlorobenzene classes. It can be used as a fluorescent dye. (See first source for structural formula.)
ELF97 phosphate is not a clinical drug and has no regulatory approvals. It is a high-purity biochemical reagent for research, specifically for histochemistry, immunohistochemistry, and flow cytometry. It is used to detect alkaline phosphatase (ALP) activity in bone, liver, and stem cell research. The water-insoluble nature of the product allows for high-resolution, localized detection, making it superior to soluble fluorogenic substrates. It is stored at -20degC, protected from light. ELF97 phosphate is also used as a substrate for ELF (Enzyme-Labeled Fluorescence) technology. |
| Molecular Formula |
C14H9CL2N2O5P
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|---|---|
| Molecular Weight |
387.11
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| Exact Mass |
385.963
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| CAS # |
147394-94-3
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| PubChem CID |
135424854
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| Appearance |
Typically exists as solids at room temperature
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| Density |
1.78g/cm3
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| Boiling Point |
654.9ºC at 760mmHg
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| Flash Point |
349.8ºC
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| Vapour Pressure |
4.84E-18mmHg at 25°C
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| Index of Refraction |
1.729
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| LogP |
3.781
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| Hydrogen Bond Donor Count |
3
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| Hydrogen Bond Acceptor Count |
6
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| Rotatable Bond Count |
3
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| Heavy Atom Count |
24
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| Complexity |
584
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| Defined Atom Stereocenter Count |
0
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| SMILES |
O=P(O)(OC1=CC=C(Cl)C=C1C(NC2=C3C=C(Cl)C=C2)=NC3=O)O
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| InChi Key |
IXZONVAEGFOVSF-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C14H9Cl2N2O5P/c15-7-1-3-11-9(5-7)14(19)18-13(17-11)10-6-8(16)2-4-12(10)23-24(20,21)22/h1-6H,(H,17,18,19)(H2,20,21,22)
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| Chemical Name |
[4-chloro-2-(6-chloro-4-oxo-3H-quinazolin-2-yl)phenyl] dihydrogen phosphate
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| Synonyms |
ELFP
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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 |
| 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) |
May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples
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| Solubility (In Vivo) |
Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.
Injection Formulations
Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)(e.g. IP/IV/IM/SC) *Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil) Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals). View More
Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)] Oral Formulations
Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium) Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals). View More
Oral Formulation 3: Dissolved in PEG400 (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.5832 mL | 12.9162 mL | 25.8325 mL | |
| 5 mM | 0.5166 mL | 2.5832 mL | 5.1665 mL | |
| 10 mM | 0.2583 mL | 1.2916 mL | 2.5832 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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