| Size | Price | Stock | Qty |
|---|---|---|---|
| 5mg |
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| 10mg |
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| 25mg | |||
| 50mg | |||
| 100mg | |||
| Other Sizes |
| Targets |
Fluorogenic substrate
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|---|---|
| ln Vitro |
At 4 hours, there was a notable rise in caspase-3 activity in the Ac-DEVD-AFC group compared to the control after 1 hour. The Photofrin and LPLI groups did not exhibit a statistically significant elevation in caspase-3 activity. When caspase-3 is activated, the disruption of Ac-DEVD-AFC can be considerably reduced by transfecting shRNA-BiML [1].
To further confirm above observation, AFC (an indicator of caspase-3 activation) release assay was applied. Activated caspase-3 but not pro-caspase-3 does exert proteolytic activation on the Ac-DEVD-AFC substrate. As shown in Fig. 3E, after incubation with Ac-DEVD-AFC for 1 hour, significant increase of caspase-3 activity was observed at 4 hour after PPT (column 5) compared with control. There were no significant increases of caspase-3 activity in Photofrin and LPLI group (column 2 and 3). The cleavage of Ac-DEVD-AFC in response to caspase-3 activation was remarkably inhibited by shRNA-BimL transfection (column 6). These results demonstrated that Bim was involved in the PPT-induced caspase-3 activity, which was consistent with the result by western blot analysis [1]. |
| Cell Assay |
Fluorometric assay for caspase-3 activity [1]
For the detection of caspase-3 activity, PBS washed cell pellets (derive from either the medium or the adherent cells) were resuspended in extract buffer [25 mM HEPES (pH7.4), 0.1% TritonX-l00, 10% glycerol, 5 mM DTT, 1mM phenylmethylsulfonyl fluoride, 10 mg/ml pepstatin, and 10 mg/ml Leupeptin] and vortexed vigorously. 20μl of extract (corresponding to 10% of the sample) were incubated with the caspase-3 fluorogenic substrates Ac-DEVD-AFC at 100 μM final concentration at room temperature, and caspase-3 activity was measured continuously by monitoring the release of fluorigenic AFC at 37°C. The excitation wavelength of AFC was 400 nm and the emission wavelength was 530 nm using auto microplate reader. As a parallel study to confirm and compare the caspase-3 detection in our experiments, batch of cells were exposed to UV irradiation, which a well-established protocol to induce apoptosis. For UV irradiation-induced apoptosis, culture medium was removed, and cells were rinsed with PBS and irradiated at fluence of 120 mJ/cm2 (253.7 nm, 200 μW/cm2), and then medium was restored. |
| References | |
| Additional Infomation |
Background/Objective: Photodynamic therapy (PDT) is a promising non-invasive technique that has been successfully applied to the treatment of human cancers. Studies have shown that Bcl-2 family proteins play an important role in PDT-induced apoptosis. However, it remains unclear whether Bcl-2-interacting cell death mediators (Bim) are involved in PDT. This study aimed to investigate the effect of Bim on photofrin-induced apoptosis in human lung adenocarcinoma ASTC-a-1 cells. Methods: Intracellular Bim/Bax translocation was observed using laser confocal scanning microscopy. Intracellular Bim protein and activated caspase-3 levels were detected by Western blot. Caspase-3 activity was determined using Caspase-3 fluorescent substrate (Ac-DEVD-AFC) assay. Apoptosis induction was detected by Hoechst 33258 and PI staining and flow cytometry. The effect of Bim on PPT-induced apoptosis was investigated using RNAi technology. Results: BimL translocated to mitochondria upon PPT stimulation, similar to the activation of the downstream pro-apoptotic protein Bax. PPT increases intracellular Bim levels and activates caspase-3, while RNAi knockdown of Bim significantly inhibits caspase-3 activity. In cells transfected with shRNA-Bim, PPT-induced apoptosis was suppressed. Conclusion: We have demonstrated that Bim is involved in PPT-induced apoptosis in human ASTC-a-1 lung adenocarcinoma cells and suggest that enhancing Bim activity may be a potential strategy for treating human cancer. [1]
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| Molecular Formula |
C30H34N5O13F3
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|---|---|
| Molecular Weight |
729.611860000001
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| Exact Mass |
729.21
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| CAS # |
201608-14-2
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| PubChem CID |
644234
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| Sequence |
N-Acetyl-Asp-Glu-Val-Asp-7-amido-4-trifluoroMethylcoumarin
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| SequenceShortening |
DEVD; Ac-DEVD-7-amido-4-trifluoroMethylcoumarin
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| Appearance |
Light yellow to yellow solid powder
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| Density |
1.5±0.1 g/cm3
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| Boiling Point |
1152.7±65.0 °C at 760 mmHg
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| Flash Point |
650.9±34.3 °C
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| Vapour Pressure |
0.0±0.3 mmHg at 25°C
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| Index of Refraction |
1.575
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| LogP |
1.58
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| Hydrogen Bond Donor Count |
8
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| Hydrogen Bond Acceptor Count |
16
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| Rotatable Bond Count |
17
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| Heavy Atom Count |
51
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| Complexity |
1430
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| Defined Atom Stereocenter Count |
4
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| SMILES |
CC(C)[C@@H](C(=O)N[C@@H](CC(=O)O)C(=O)NC1=CC2=C(C=C1)C(=CC(=O)O2)C(F)(F)F)NC(=O)[C@H](CCC(=O)O)NC(=O)[C@H](CC(=O)O)NC(=O)C
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| InChi Key |
GZDRODOYEFEHGG-NUDCOPPTSA-N
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| InChi Code |
InChI=1S/C30H34F3N5O13/c1-12(2)25(38-26(47)17(6-7-21(40)41)36-28(49)18(10-22(42)43)34-13(3)39)29(50)37-19(11-23(44)45)27(48)35-14-4-5-15-16(30(31,32)33)9-24(46)51-20(15)8-14/h4-5,8-9,12,17-19,25H,6-7,10-11H2,1-3H3,(H,34,39)(H,35,48)(H,36,49)(H,37,50)(H,38,47)(H,40,41)(H,42,43)(H,44,45)/t17-,18-,19-,25-/m0/s1
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| Chemical Name |
(4S)-4-[[(2S)-2-acetamido-3-carboxypropanoyl]amino]-5-[[(2S)-1-[[(2S)-3-carboxy-1-oxo-1-[[2-oxo-4-(trifluoromethyl)chromen-7-yl]amino]propan-2-yl]amino]-3-methyl-1-oxobutan-2-yl]amino]-5-oxopentanoic acid
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| Synonyms |
Ac-DEVD-AFC; Ac-Asp-Glu-Val-Asp-AFC; 201608-14-2; MFCD01310970; N-Acetyl-Asp-Glu-Val-Asp-7-amido-4-trifluoromethylcoumarin; (4S)-4-[[(2S)-2-acetamido-3-carboxypropanoyl]amino]-5-[[(2S)-1-[[(2S)-3-carboxy-1-oxo-1-[[2-oxo-4-(trifluoromethyl)chromen-7-yl]amino]propan-2-yl]amino]-3-methyl-1-oxobutan-2-yl]amino]-5-oxopentanoic acid; Ac-Asp-Glu-Val-Asp-7-amino-4-trifluoromethylcoumarin; SCHEMBL1180114;
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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: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture and light. |
| 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 : ≥ 50 mg/mL (~68.53 mM)
H2O : ~0.67 mg/mL (~0.92 mM) |
|---|---|
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (3.43 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 (3.43 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in 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 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: ≥ 2.5 mg/mL (3.43 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 | 1.3706 mL | 6.8530 mL | 13.7060 mL | |
| 5 mM | 0.2741 mL | 1.3706 mL | 2.7412 mL | |
| 10 mM | 0.1371 mL | 0.6853 mL | 1.3706 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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