DR probe
DR probe is a "dual-key-and-lock" fluorescent probe designed based on the Resorufin (HY-123533) scaffold (Ex/Em = 647 nm/663-738 nm). DR probe can be sequentially activated by hydrogen peroxide to form the intermediate SR, which acts as a tyrosinase substrate to trigger a fluorescence turn-on signal. DR probe can distinguish normal melanocytes from melanoma cells. DR probe supports cell imaging and can be applied to mouse melanoma models to achieve melanoma diagnosis with higher accuracy and lower false-positive rates. DR probe is applicable to relevant research on melanoma.
For research use only. We do not sell to patients.
- Formula: C32H30BNO6
- Molecular Weight:535.39
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Storage:
Please store the product under the recommended conditions in the Certificate of Analysis.
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Biological Activity
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Chemical Information
- Purity & Documentation
- References
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Help & FAQs
Biological Activity
The DR probe (5-20 μM; 90 min-3.5 h) exhibits a 56-fold fluorescence turn-on effect at 588 nm only when sequentially activated by H2O2 and TYR under physiological conditions, with a limit of detection of 28.1 nM for the H2O2-mediated activation process[1].
DR probe (0-30 μM; 1-24 h) shows no cytotoxicity to HSF, PIG1 and A375 cells even at the maximum tested concentration of 30 μM, and it can selectively produce strong fluorescence in A375 melanoma cells (8-fold higher than that in normal PIG1 melanocytes) by responding to endogenous H2O2 and TYR[1].
Guidelines (The following is our recommended protocol, which serves only as a guide and should be modified according to your specific needs).
Cell Fluorescence Imaging:
1.\tCell Preparation: Seed human malignant melanoma cells (A375), normal melanocytes (PIG1) or skin fibroblasts (HSF) into confocal culture dishes, and culture them routinely.
(Optional Control Group): Pre-treat cells with Kojic acid (HY-W050154) (200 μM, a TYR inhibitor) or NAC (HY-B0215) (200 μM, an H2O2 scavenger) for 1 hour.
2.\tProbe Incubation:
Aspirate the culture medium, and add serum-free medium or PBS containing 10 μM DR (or SR as a control).
Incubate the cells at 37°C in a 5% CO2 atmosphere for 3.5 hours.
3.\tWashing: Aspirate the probe solution, and wash the cells 3 times with PBS to remove unbound probes.
4.\tMicroscopic Imaging:
Use a laser confocal scanning microscope.
Excitation (Ex) = 561 nm
Emission (Em) = 570-700 nm (to detect the near-infrared fluorescence of released Resorufin).
Expected Results: A375 cells show strong red fluorescence; PIG1/HSF cells and the inhibitor/NAC treatment groups exhibit extremely low fluorescence.
MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.
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Cell Line:human skin fibroblast (HSF) cells, normal human melanocyte (PIG1) cells, human malignant melanoma (A375) cells
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Concentration:0, 2.5, 5, 10, 20, 30 μM (cytotoxicity assay); 10 μM (imaging; inhibition studies with 200 μM kojic acid/NAC pre-treatment)
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Incubation Time:24 h (cytotoxicity assay); 1, 2, or 3.5 h (imaging); 1 h (kojic acid/NAC pre-treatment) then 3.5 h (DR incubation, inhibition studies)
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Result:Resulted in > 80% cell survival in HSF, PIG1, and A375 cells at 30 μM.
Caused progressive fluorescence intensity increases over 1, 2, and 3.5 h in A375 cells, reaching a 13-fold increase.
Produced virtually undetectable fluorescence in HSF and PIG1 cells, while showing strong fluorescence (8-fold higher intensity than PIG1 cells) in A375 cells.
Showed near-basal fluorescence levels in A375 cells pre-treated with kojic acid or NAC.
Guidelines (The following is our recommended protocol, which serves only as a guideline and should be modified according to your specific needs).
Mouse Live-Body Melanoma Imaging1. Animal Model: 4-week-old Balb/c nude mice.
Subcutaneously inoculate A375 cells (e.g., 1×107 cells per mouse, in the axillary region) and wait for tumor growth for 7-21 days.
(Optional control group): Pre-treat tumor-bearing mice with 1 mM Kojic acid or 1 mM NAC via tail vein injection for 1 hour.
2. Probe Administration and Imaging:
Inject DR solution (200 μM, at an appropriate volume such as 100 μL or adjusted based on body weight) via tail vein.
Anesthetize the mice and perform in vivo fluorescence imaging at 30, 60, 120, and 180 minutes post-injection.
Detection parameters: Refer to cell experiments for excitation/emission windows.
Expected results: The fluorescence intensity at the tumor site reaches its peak at approximately 120 minutes (about 46-fold higher than that of the control group), and the signal is significantly blocked in the inhibitor group.
MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.
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Animal Model:Balb/c nude (4-week-old female)[1]
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Dosage:200 μM
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Administration:i.v. (tail vein); single injection
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Result:Reached peak fluorescence signal at 120 minutes post-injection, with intensity approximately 46-fold higher than the control group in 7-day tumor xenografts.
Increased fluorescence intensity at the tumor site with longer tumor growth times (7, 10, 14, 21 days).
Decreased fluorescence intensity by 180 minutes post-injection.\nExhibited a tumor fluorescence signal only ~6.6-fold higher than the control group in mice pretreated with N-acetyl-L-cysteine, which was significantly lower than the signal in unpretreated xenograft mice.
Showed a similar reduction in fluorescence intensity in mice pretreated with kojic acid.\nProduced significant fluorescence signals 2 hours post-injection in both small ( < 1 mm3) and large (16 mm3) 14-day xenograft tumors.
Detected strong fluorescence in lung tissue of metastatic melanoma mice, alongside high tyrosinase levels in lung and tumor tissues (confirmed via Western blot).
Chemical Information
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Molecular Weight 535.39
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Formula C32H30BNO6
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SMILES
CC1(OB(OC1(C)C)C2=CC=C(C=C2)COC3=CC(COC4=CC=C5N=C6C=CC(C=C6OC5=C4)=O)=CC=C3)C
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Shipping
Room temperature in continental US; may vary elsewhere.
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Storage
Please store the product under the recommended conditions in the Certificate of Analysis.
Purity & Documentation
References
Calculators
Concentration (start) × Volume (start) = Concentration (final) × Volume (final)
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