Fura Red is a Ca2+-sensitive fluorescent dye whose fluorescence decreases as the [Ca2+] concentration increases.

Physicochemical Properties

Molecular Formula	C41H44N4O20S
Molecular Weight	944.87
Exact Mass	944.227
CAS #	149732-62-7
PubChem CID	6438360
Appearance	Typically exists as solid at room temperature
Density	1.48g/cm3
Index of Refraction	1.624
LogP	1.535
Hydrogen Bond Donor Count	2
Hydrogen Bond Acceptor Count	23
Rotatable Bond Count	32
Heavy Atom Count	66
Complexity	1750
Defined Atom Stereocenter Count	0
SMILES	CC(OCOC(CN(C1C=CC(C)=CC=1OCCOC1C=C2C=C(OC2=CC=1N(CC(OCOC(=O)C)=O)CC(OCOC(=O)C)=O)/C=C1\NC(=S)NC\1=O)CC(OCOC(=O)C)=O)=O)=O
InChi Key	MCEXQZRGUKALLT-VVEOGCPPSA-N
InChi Code	InChI=1S/C41H44N4O20S/c1-23-6-7-31(44(15-36(50)61-19-57-24(2)46)16-37(51)62-20-58-25(3)47)34(10-23)55-8-9-56-35-12-28-11-29(13-30-40(54)43-41(66)42-30)65-33(28)14-32(35)45(17-38(52)63-21-59-26(4)48)18-39(53)64-22-60-27(5)49/h6-7,10-14H,8-9,15-22H2,1-5H3,(H2,42,43,54,66)/b30-13+
Chemical Name	acetyloxymethyl 2-[N-[2-(acetyloxymethoxy)-2-oxoethyl]-2-[2-[[6-[bis[2-(acetyloxymethoxy)-2-oxoethyl]amino]-2-[(E)-(5-oxo-2-sulfanylideneimidazolidin-4-ylidene)methyl]-1-benzofuran-5-yl]oxy]ethoxy]-4-methylanilino]acetate
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
Shipping Condition	Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Biological Activity

ln Vitro	Guidelines[2] (The protocol listed below is our suggested one; it should be adjusted to meet your unique requirements as it just serves as a guide). Cell Labeling: 1. Proceed with the regular protocol for incubating the cells. 2. In 37°C Hanks Balanced Salt Solution (HBSS) with 1 μM Fura Red, AM (unless otherwise noted) and 0.01% Pluronic F127, suspend cells at 1x107 cells/mL. Incubate for 30 minutes in a 37°C water bath. 3. Re-suspend cells to 1x107 cells/mL in HEPES Buffered Saline Solution using viability dye, SYTOX Green, after washing them in HEPES Buffered Saline Solution (HBSS with 1 mM CaCl2, 0.5 mM MgCl2, 0.1% BSA, and 10 mM HEPES). 4. Cells should equilibrate in a water bath at 37°C for at least ten minutes. 5. Use a flow cytometer, fluorescence microscopy, or fluorescence microplate reader to analyze the sample. Fura Red is measured using ratiometric analysis using two lasers: the green laser (532 nm) and the violet laser (406 nm). Two distinct filter sets are used to detect emission: the Violet laser (630LP and 660/20 BP) is used to monitor increases in emission, while the Green laser (685LP and 710/50 BP) is used to detect decreases in emission.
References	[1]. IgG-induced Ca2+ oscillations in differentiated U937 cells; a study using laser scanning confocal microscopy and co-loaded fluo-3 and fura-red fluorescent probes. Cell Calcium. 1995 Nov;18(5):377-89. [2]. Ratiometric analysis of fura red by flow cytometry: a technique for monitoring intracellular calcium flux in primary cell subsets. PLoS One. 2015 Apr 2;10(3):e0119532.
Additional Infomation	Fura red is a 1-benzofuran substituted at position 2 by a (5-oxo-2-thioxoimidazolidin-4-ylidene)methyl group, and at C-5 and C-6 by heavily substituted oxygen and nitrogen functionalities respectively. It has a role as a fluorochrome.

Solubility Data

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

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 (e.g. IP/IV/IM/SC) Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline) *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). 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)] *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. Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline) Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline) Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline) Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil) Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL 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). Oral Formulation 3: Dissolved in PEG400 Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose Oral Formulation 6: Mixing with food powders Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	1.0583 mL	5.2917 mL	10.5835 mL
	5 mM	0.2117 mL	1.0583 mL	2.1167 mL
	10 mM	0.1058 mL	0.5292 mL	1.0583 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.