JC-1 does not have a specific pharmacological target. It is a fluorescent probe used for research applications rather than a therapeutic agent. JC-1 specifically labels mitochondria in living cells and its fluorescence emission shifts from green (monomeric form) to orange (J-aggregate form) as mitochondrial membrane potential increases. This property makes JC-1 a valuable tool for studying mitochondrial function, apoptosis, and cellular energy metabolism. The probe is also used for testing P-glycoprotein (Pgp) activity.

JC-1 staining solution composition 1.1 Make the stock solution with DMSO to achieve a concentration of 5 mg/mL of JC-1. For instance, dissolve 5 mg of JC-1 in 1 mL of DMSO. It is advised to aliquot the JC-1 storage solution and store it in the dark at -20°C or -80°C. 1.2 Working solution replacement: Make a JC-1 working solution at a concentration of 1–20 μg/mL by mixing PBS or previously prepared serum-free cell bone marrow with the storage solution. Note: 1) Please utilize the existing configuration and modify the JC-1's working liquid concentration in accordance with the real circumstances. 2) You can add 20% diluent Pluronic F127 solution to the working solution, with a final concentration of 0.02-0.05%, if the effect of JC-1 entering the cells is not good. Pluronic F127 can assist JC-1 in entering the cell by preventing it from aggregating in the buffer. JC-1 staining 1) Plate cells at a density of 5×105 cells/mL using a 6-well plate as an example. In an incubator with 5% CO2 and 37°C, cultivate overnight. Note: During induction, it is advised that the cell density not surpass 1×106/mL. Additionally, you can change the density to a density that suits your own culture type. 2) Once sterile, take 0.5 mL of the cell suspension. 3) Centrifuge for three to five minutes at 400 g; remove supernatant. 4) After resuspending the cells in 1 milliliter of JC-1 working solution, incubate for 15 to 30 minutes on the side wall of an incubator set at 37°C with 5% CO2. 5) After that, 6) resuspend the cells in 2 milliliters of buffer or cell culture media, and centrifuge for an additional 5 minutes at 400 g. Discard the supernatant and repeat twice. 7) Re-suspend the cells in 1 mL of new culture medium or buffer, and then go straight to the observation of fluorescence microscopy or flow cytometry analysis. 8) Data analysis (flow cytometry): Red JC-1 aggregates from healthy cell mitochondria are detected using the FL2 channel; green ones containing green Note: If the cells are to be used for microplate reader detection, resuspend them with 300 μL buffer and proceed as directed. Transfer the stained cells into a 100 μL volume of a light-tight 96-well plate, and then carry out fluorescence microplate analysis.
In vitro, JC-1 is used as a probe for measuring mitochondrial membrane potential by flow cytometry. The dye accumulates in mitochondria in a membrane potential-dependent manner. At low membrane potential, JC-1 exists as monomers emitting green fluorescence (529 nm). At high membrane potential, JC-1 forms J-aggregates emitting orange-red fluorescence. The ratio of red to green fluorescence provides a quantitative measure of mitochondrial membrane potential. JC-1 is also used to test Pgp activity.

JC-1 is not used as a therapeutic agent and therefore in vivo pharmacological activity is not applicable. The probe is used in research applications for assessing mitochondrial function in cells and tissues. It can be used to evaluate mitochondrial membrane potential changes in response to various stimuli, including apoptosis-inducing agents, metabolic modulators, and mitochondrial toxins. However, JC-1 is not administered in vivo as a therapeutic compound.

For JC-1, standard experimental protocols involve loading cells with JC-1 dye (typically 2-10 µM) for 15-30 minutes at 37°C. After washing, cells are analyzed by flow cytometry, fluorescence microscopy, or fluorescence spectroscopy. The ratio of red (J-aggregates, excitation 550 nm, emission 590 nm) to green (monomers, excitation 488 nm, emission 529 nm) fluorescence is calculated to determine mitochondrial membrane potential.

For JC-1, standard cellular assays involve treatment of cells with the test compound or experimental condition followed by JC-1 staining. Cells are incubated with JC-1, washed, and analyzed by flow cytometry or fluorescence microscopy. Changes in mitochondrial membrane potential are detected as shifts in the red/green fluorescence ratio. This assay is commonly used to assess mitochondrial dysfunction in apoptosis, drug toxicity, and metabolic studies.

JC-1 is not used in standard in vivo animal models as a therapeutic agent. It is a research tool for assessing mitochondrial function in isolated cells or tissues. For in vivo imaging applications, JC-1 may be administered locally or systemically for short-term imaging studies, but this is not a standard pharmacological efficacy protocol.

JC-1 has a molecular weight of 652.22-652.23 and formula C25H27Cl4IN4. It appears as a dark violet powder with a purity of ≥95-98%. The compound has a melting point of 275-278°C. It is soluble in DMSO at 5 mg/mL and should be stored at room temperature.

JC-1 is a research tool and not a therapeutic agent. As a fluorescent dye, it is not intended for human therapeutic use. The compound should be handled with standard laboratory safety precautions. Comprehensive toxicological evaluation for therapeutic applications is not applicable.

[1]. JC-1: alternative excitation wavelengths facilitate mitochondrial membrane potential cytometry. Cell Death Dis. 2012 Nov 22;3:e430.

[2]. Ratiometric high-resolution imaging of JC-1 fluorescence reveals the subcellular heterogeneity of astrocytic mitochondria. Pflügers Archiv - European Journal of Physiology. 2011,462(5): 693-708.

[3]. Real-time analysis of amyloid fibril formation of α-synuclein using a fibrillation-state-specific fluorescent probe of JC-1. Biochem. J. 2009, 418:311-323.

[4]. JC-1, but not DiOC6(3) or rhodamine 123, is a reliable fluorescent probe to assess delta psi changes in intact cells: implications for studies on mitochondrial functionality during apoptosis. FEBS Lett. 1997 Jul 7;411(1):77-82.

1,1',3,3'-Tetraethyl-5,5',6,6'-Tetrachloroimidazole carbocyanine iodide is a cyanine dye and an organic iodide salt. It is used as a fluorescent dye. It contains 1,1',3,3'-tetraethyl-5,5',6,6'-tetrachloroimidazole carbocyanine.
JC-1 is also known as 5,5',6,6'-Tetrachloro-1,1',3,3'-tetraethylbenzimidazolocarbocyanine iodide. It is a cationic, lipophilic fluorescent dye widely used to assess mitochondrial membrane potential (ΔΨm) in live cells. The dye's fluorescence color changes with mitochondrial transmembrane potential. JC-1 is also used for testing Pgp activity. No drug development or clinical trial status applies.

C25H27CL4IN4

652.2235

650.003

3520-43-2

3520-43-2;

5492929

Purple to purplish red solid powder

4.157

0

3

6

34

625

0

CCN1C2=CC(=C(C=C2[N+](=C1/C=C/C=C3N(C4=CC(=C(C=C4N3CC)Cl)Cl)CC)CC)Cl)Cl.[I-]

FYNNIUVBDKICAX-UHFFFAOYSA-M

InChI=1S/C25H27Cl4N4.HI/c1-5-30-20-12-16(26)17(27)13-21(20)31(6-2)24(30)10-9-11-25-32(7-3)22-14-18(28)19(29)15-23(22)33(25)8-4/h9-15H,5-8H2,1-4H31H/q+1/p-1

1H-Benzimidazolium, 5,6-dichloro-2-(3-(5,6-dichloro-1,3-diethyl-1,3-dihydro-2H-benzimidazol-2-ylidene)-1-propenyl)-1,3-diethyl-, iodide

JC 1 JC-1 JC1

2934.99.9001

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.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~5 mg/mL (~7.67 mM)
H2O : < 0.1 mg/mL

Solubility in Formulation 1: 1.25 mg/mL (1.92 mM) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), suspension solution; with sonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 12.5 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.

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Solubility in Formulation 2: 1.25 mg/mL (1.92 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 12.5 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.

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 (Please use freshly prepared in vivo formulations for optimal results.)