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  4. Fluorescein Diacetate

Fluorescein Diacetate  (Synonyms: 3,6-Diacetoxyfluoran; Di-O-acetylfluorescein)

Cat. No.: HY-D0719 Purity: 99.87%
COA Handling Instructions

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Fluorescein diacetate is a cell permeable esterase-substrate. Fluorescein diacetate can be used as a fluorogenic substrate for hGSTP1-1.

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Fluorescein Diacetate Chemical Structure

Fluorescein Diacetate Chemical Structure

CAS No. : 596-09-8

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Based on 2 publication(s) in Google Scholar

Fluorescein Diacetate Related Antibodies

Top Publications Citing Use of Products

  • Biological Activity

  • Protocol

  • Purity & Documentation

  • References

  • Customer Review

Description

Fluorescein diacetate is a cell permeable esterase-substrate. Fluorescein diacetate can be used as a fluorogenic substrate for hGSTP1-1.

In Vitro

Fluorescein diacetate (FDA) is an acetylated derivative of the green fluorescent dye fluorescein[1]. Fluorescein diacetate (FDA), a fluorescent probe used for vital staining, is a fluorescently activated by esterolytic activity of human Pi-class glutathione S-transferase (hGSTP1) selectively among various cytosolic GSTs. Fluorescence activation of Fluorescein diacetate susceptible to GST inhibitors is observed in MCF7 cells exogenously overexpressing hGSTP1, but not in cells overexpressing hGSTA1 or hGSTM1. Fluorescein diacetate can be used as a fluorogenic substrate for hGSTP1-1. To investigate whether the fluorescence activation is due to hGSTP1 activity, Fluorescein diacetate is incubated with recombinant hGSTP1-1 and GSH in vitro. Remarkable fluorescence activation is observed in the presence of both hGSTP1-1 and GSH, whereas only slight activation is observed in the absence of either of them or when the enzyme is heat inactivated. This suggests that the fluorescence activation of Fluorescein diacetate depends on hGSTP1-1 activity. From the linear relationship between the rate of increase in fluorescence and the hGSTP1-1 concentration, the specific activity of the enzyme for 1 μM Fluorescein diacetate is determined to be 79±15 nmol/min/mg protein. Fluorescein diacetate is applicable as a fluorogenic substrate for evaluating inhibitors of GSTP1-1 in vitro. For Fluorescein diacetate as a substrate, both Ethacrynic acid (EA) and NBDHEX suppress the hGSTP1-1-dependent fluorescent increase in a concentration-dependent manner, with IC50s of 3.3±0.5 μM and 0.61±0.04 μM, respectively[2].

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.

Fluorescein Diacetate Related Antibodies
Molecular Weight

416.38

Formula

C24H16O7
CAS No.
Appearance

Solid

Color

Light yellow to yellow

SMILES

O=C1OC2(C3=C(OC4=C2C=CC(OC(C)=O)=C4)C=C(OC(C)=O)C=C3)C5=C1C=CC=C5
Shipping

Room temperature in continental US; may vary elsewhere.
Storage

-20°C, protect from light

*In solvent : -80°C, 6 months; -20°C, 1 month (protect from light)

Solvent & Solubility
In Vitro: 

DMSO : 26.67 mg/mL (64.05 mM; Need ultrasonic; Hygroscopic DMSO has a significant impact on the solubility of product, please use newly opened DMSO)

Preparing
Stock Solutions
Concentration Solvent Mass 1 mg 5 mg 10 mg
1 mM 2.4017 mL 12.0083 mL 24.0165 mL
5 mM 0.4803 mL 2.4017 mL 4.8033 mL
View the Complete Stock Solution Preparation Table

* Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month (protect from light). When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.

  • Molarity Calculator

  • Dilution Calculator

Mass (g) = Concentration (mol/L) × Volume (L) × Molecular Weight (g/mol)

Mass
=
Concentration
×
Volume
×
Molecular Weight *

Concentration (start) × Volume (start) = Concentration (final) × Volume (final)

This equation is commonly abbreviated as: C1V1 = C2V2

Concentration (start)

C1

×
Volume (start)

V1

=
Concentration (final)

C2

×
Volume (final)

V2

In Vivo:

Select the appropriate dissolution method based on your experimental animal and administration route.

For the following dissolution methods, please ensure to first prepare a clear stock solution using an In Vitro approach and then sequentially add co-solvents:
To ensure reliable experimental results, the clarified stock solution can be appropriately stored based on storage conditions. As for the working solution for in vivo experiments, it is recommended to prepare freshly and use it on the same day.
The percentages shown for the solvents indicate their volumetric ratio in the final prepared solution. If precipitation or phase separation occurs during preparation, heat and/or sonication can be used to aid dissolution.

  • Protocol 1

    Add each solvent one by one:  10% DMSO    90% (20% SBE-β-CD in Saline)

    Solubility: ≥ 2.08 mg/mL (5.00 mM); Clear solution

    This protocol yields a clear solution of ≥ 2.08 mg/mL (saturation unknown).

    Taking 1 mL working solution as an example, add 100 μL DMSO stock solution (20.8 mg/mL) to 900 μL 20% SBE-β-CD in Saline, and mix evenly.

    Preparation of 20% SBE-β-CD in Saline (4°C, storage for one week): 2 g SBE-β-CD powder is dissolved in 10 mL Saline, completely dissolve until clear.
  • Protocol 2

    Add each solvent one by one:  10% DMSO    90% Corn Oil

    Solubility: ≥ 2.08 mg/mL (5.00 mM); Clear solution

    This protocol yields a clear solution of ≥ 2.08 mg/mL (saturation unknown). If the continuous dosing period exceeds half a month, please choose this protocol carefully.

    Taking 1 mL working solution as an example, add 100 μL DMSO stock solution (20.8 mg/mL) to 900 μL Corn oil, and mix evenly.

In Vivo Dissolution Calculator
Please enter the basic information of animal experiments:

Dosage

mg/kg

Animal weight
(per animal)

g

Dosing volume
(per animal)

μL

Number of animals

Recommended: Prepare an additional quantity of animals to account for potential losses during experiments.
Please enter your animal formula composition:
%
DMSO +
% 
+
%
Tween-80 +
%
Saline
Recommended: Keep the proportion of DMSO in working solution below 2% if your animal is weak.
The co-solvents required include: DMSO, . All of co-solvents are available by MedChemExpress (MCE). , Tween 80. All of co-solvents are available by MedChemExpress (MCE).
Calculation results:
Working solution concentration: mg/mL
Method for preparing stock solution: mg drug dissolved in μL  DMSO (Stock solution concentration: mg/mL).

*In solvent : -80°C, 6 months; -20°C, 1 month (protect from light)

The concentration of the stock solution you require exceeds the measured solubility. The following solution is for reference only. If necessary, please contact MedChemExpress (MCE).
Method for preparing in vivo working solution for animal experiments: Take μL DMSO stock solution, add μL . μL , mix evenly, next add μL Tween 80, mix evenly, then add μL Saline.
 If the continuous dosing period exceeds half a month, please choose this protocol carefully.
Please ensure that the stock solution in the first step is dissolved to a clear state, and add co-solvents in sequence. You can use ultrasonic heating (ultrasonic cleaner, recommended frequency 20-40 kHz), vortexing, etc. to assist dissolution.
Purity & Documentation

Purity: 99.87%

COA (252 KB) HNMR (269 KB) LCMS (268 KB)

Handling Instructions (2659 KB)
References
Cell Assay
[2]

MCF7 cells (2-3×105) are seeded in a 35-mm glass bottom dish before the experiment. Prior to imaging, cells are washed with 1 mL PBS, then incubated in 1 mL Hanks’ Balanced Salt solution (HBSS(+) without phenol red) containing 1 μM Fluorescein diacetate (0.1% DMSO as a cosolvent) for 5 min at 37°C. Cells are washed twice with 1 mL PBS and 1 mL HBSS is added before imaging. Fluorescence images of fluorescein and DsRed-Express2 were acquired in the FITC channel (excitation at 473 nm) and the DsRed channel (excitation 559 nm). The 16-bit images obtained are analysed[2].

MCE has not independently confirmed the accuracy of these methods. They are for reference only.
References

Complete Stock Solution Preparation Table

* Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month (protect from light). When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.

Optional Solvent Concentration Solvent Mass 1 mg 5 mg 10 mg 25 mg
DMSO 1 mM 2.4017 mL 12.0083 mL 24.0165 mL 60.0413 mL
5 mM 0.4803 mL 2.4017 mL 4.8033 mL 12.0083 mL
10 mM 0.2402 mL 1.2008 mL 2.4017 mL 6.0041 mL
15 mM 0.1601 mL 0.8006 mL 1.6011 mL 4.0028 mL
20 mM 0.1201 mL 0.6004 mL 1.2008 mL 3.0021 mL
25 mM 0.0961 mL 0.4803 mL 0.9607 mL 2.4017 mL
30 mM 0.0801 mL 0.4003 mL 0.8006 mL 2.0014 mL
40 mM 0.0600 mL 0.3002 mL 0.6004 mL 1.5010 mL
50 mM 0.0480 mL 0.2402 mL 0.4803 mL 1.2008 mL
60 mM 0.0400 mL 0.2001 mL 0.4003 mL 1.0007 mL
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Fluorescein Diacetate Related Classifications

Help & FAQs
  • Do most proteins show cross-species activity?

    Species cross-reactivity must be investigated individually for each product. Many human cytokines will produce a nice response in mouse cell lines, and many mouse proteins will show activity on human cells. Other proteins may have a lower specific activity when used in the opposite species.

Keywords:

Fluorescein Diacetate596-09-83,6-Diacetoxyfluoran Di-O-acetylfluoresceinBiochemical Assay ReagentsInhibitorinhibitorinhibit

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