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Evans Blue

Alias: Direct Blue 53
Cat No.:V2710 Purity: ≥98%
Evans Blue (Direct Blue 53)is a potent inhibitor of the uptake of L-glutamate into synaptic vesicles, it is also an AMPA/kainate receptor antagonist.
Evans Blue
Evans Blue Chemical Structure CAS No.: 314-13-6
Product category: GluR
This product is for research use only, not for human use. We do not sell to patients.
Size Price Stock Qty
500mg
2g
5g
10g
25g
50g
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Purity & Quality Control Documentation

Purity: ≥98%

Product Description

Evans Blue (Direct Blue 53) is a potent inhibitor of the uptake of L-glutamate into synaptic vesicles, it is also an AMPA/kainate receptor antagonist. Evans Blue inhibits DNA binding of NF-κB at a low concentration of 100 μM. It has proven over the years to be a dependable stain for microscopic determination of cell death. Evans Blue is also a known blocker of a subset of a-amino-3-hydroxy-5-methyl-isoxazole/kainate receptors (IC50=355 nM) for the subunit combination GluR1,2. Evans blue is the first known δ-subunit-specific antagonist of ENaC and activates large-conductance Ca2+-activated K+ channels in sheep bladder myocytes and cultured endothelial cells of human umbilical veins.

Biological Activity I Assay Protocols (From Reference)
ln Vitro
Evans Blue is a disazo dye, meaning it doesn't penetrate. Evans Blue penetrates the cytoplasm and nucleus when the plasma membrane is damaged, giving them a blue stain. Cell viability can be examined using Evans Blue[1].
ln Vivo
Regardless of the stain administration route, mice's brains accumulated the same amount of Evans Blue (2% solution; 4 mL/kg) stain following intracerebral hemorrhage caused by collagenase injection [2].
Animal Protocol
Animal/Disease Models: CD-1 mice bearing intracerebral hemorrhage (ICH)[2]
Doses: 2% solution; 4 mL/kg
Route of Administration: Jugular vein or intraperitoneal (ip)injection; once
Experimental Results: demonstrated the amount of stain accumulated in the brains.
References

[1]. Evans Blue Dye: A Revisit of Its Applications in Biomedicine. Contrast Media Mol Imaging. 2018 Apr 22:2018:7628037.

[2]. Comparison Evans Blue injection routes: Intravenous versus intraperitoneal, for measurement of blood-brain barrier in a mice hemorrhage model. J Neurosci Methods. 2011 Feb 15;195(2):206-10.

Additional Infomation
Evans blue is an organic sodium salt that is the tetrasodium salt of 6,6'-{(3,3'-dimethyl[1,1'-biphenyl]-4,4'-diyl)bis[diazene-2,1-diyl]}bis(4-amino-5-hydroxynaphthalene-1,3-disulfonate). It is sometimes used as a counterstain, especially in fluorescent methods to suppress background autofluorescence. It has a role as a histological dye, a fluorochrome, a teratogenic agent and a sodium channel blocker. It contains an Evans blue(4-).
An azo dye used in blood volume and cardiac output measurement by the dye dilution method. It is very soluble, strongly bound to plasma albumin, and disappears very slowly.
See also: Evans Blue (annotation moved to).
These protocols are for reference only. InvivoChem does not independently validate these methods.
Physicochemical Properties
Molecular Formula
C34H24N6O14S4.4NA
Molecular Weight
960.81
Exact Mass
959.982
CAS #
314-13-6
Related CAS #
314-13-6
PubChem CID
9409
Appearance
White to off-white solid powder
LogP
10.785
Hydrogen Bond Donor Count
4
Hydrogen Bond Acceptor Count
20
Rotatable Bond Count
5
Heavy Atom Count
62
Complexity
1790
Defined Atom Stereocenter Count
0
InChi Key
ATNOAWAQFYGAOY-UHFFFAOYSA-J
InChi Code
InChI=1S/C34H28N6O14S4.4Na/c1-15-11-17(3-7-21(15)37-39-23-9-5-19-25(55(43,44)45)13-27(57(49,50)51)31(35)29(19)33(23)41)18-4-8-22(16(2)12-18)38-40-24-10-6-20-26(56(46,47)48)14-28(58(52,53)54)32(36)30(20)34(24)42;;;;/h3-14,41-42H,35-36H2,1-2H3,(H,43,44,45)(H,46,47,48)(H,49,50,51)(H,52,53,54);;;;/q;4*+1/p-4
Chemical Name
tetrasodium;4-amino-6-[[4-[4-[(8-amino-1-hydroxy-5,7-disulfonatonaphthalen-2-yl)diazenyl]-3-methylphenyl]-2-methylphenyl]diazenyl]-5-hydroxynaphthalene-1,3-disulfonate
Synonyms
Direct Blue 53
HS Tariff Code
2934.99.9001
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, avoid exposure to moisture.
Shipping Condition
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
Solubility Data
Solubility (In Vitro)
DMSO:100 mg/mL (104.1 mM)
Water:100 mg/mL (104.1 mM)
Ethanol:<1 mg/mL
Solubility (In Vivo)
Solubility in Formulation 1: ≥ 2.5 mg/mL (2.60 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 (2.60 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 saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 3: 2.5 mg/mL (2.60 mM) in 10% DMSO + 90% Corn Oil (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 corn oil and mix evenly.


 (Please use freshly prepared in vivo formulations for optimal results.)
Preparing Stock Solutions 1 mg 5 mg 10 mg
1 mM 1.0408 mL 5.2039 mL 10.4079 mL
5 mM 0.2082 mL 1.0408 mL 2.0816 mL
10 mM 0.1041 mL 0.5204 mL 1.0408 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.

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What is the mass of compound required to make a 10 mM stock solution in 5 ml of DMSO given that the molecular weight of the compound is 350.26 g/mol?
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  • The answer of 17.513 mg appears in the Mass box. In a similar way, you may calculate the volume and concentration.

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What volume of a given 10 mM stock solution is required to make 25 ml of a 25 μM solution?
Using the equation C1V1 = C2V2, where C1=10 mM, C2=25 μM, V2=25 ml and V1 is the unknown:
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Note: Chemical formula is case sensitive: C12H18N3O4  c12h18n3o4
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In vivo Formulation Calculator (Clear solution)
Step 1: Enter information below (Recommended: An additional animal to make allowance for loss during the experiment)
Step 2: Enter in vivo formulation (This is only a calculator, not the exact formulation for a specific product. Please contact us first if there is no in vivo formulation in the solubility section.)
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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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