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Physicochemical Properties
| Appearance | Off-white to light brown solid powder |
| Synonyms | Vari Fluor 405 SE; Vari Fluor-405 SE; |
| 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
| Targets | Fluorescent Dye |
| ln Vitro |
I. Reagent Preparation 1. Protein Preparation Requirements: • Optimal concentration: 2 mg/mL (range: 2-10 mg/mL) • pH adjustment: Maintain 8.5±0.5 using 1M sodium bicarbonate if pH<8.0 • Buffer compatibility: Must be free of primary amines (Tris/glycine) and ammonium ions 2. Dye Solution Preparation: • Dissolve VF dye in anhydrous DMSO to 10 mg/mL • Mix thoroughly using vortex or glass rod • Storage: Aliquot and protect from light at -20°C/-80°C 3. Dye Volume Calculation (Example for IgG): Given: 500μL of 2mg/mL IgG (MW=150kDa), 1mg VF488 (MW=834) Calculation steps: 1. IgG mmol = (2mg/mL×0.5mL)/150,000 = 6.7×10⁻⁶ mmol 2. VF488 mmol = 6.7×10⁻⁶ ×10 = 6.7×10⁻⁵ mmol 3. VF488 μL = (6.7×10⁻⁵×834)/0.01 = 5.6μL II. Labeling Procedure 1. Conjugation Reaction: • Slowly add calculated VF dye to protein solution • Gentle mixing followed by brief centrifugation • Incubate at RT for 60min with periodic inversion (every 10-15min) 2. Purification (Sephadex G-25): • Prepare column per manufacturer's instructions • Load reaction mixture onto column • Elute with PBS (pH7.2-7.4) • Collect conjugated protein fractions Storage: -20°C protected from light Key Considerations: 1. Light/moisture sensitivity: Prepare dye fresh for each use 2. Compatible preservatives: ≤3mM NaN₃ or ≤0.02mM thimerosal 3. Incompatible components: Glycerol (20-50%), amine-containing buffers 4. Safety: For research use only with proper PPE |
| References |
[1]. Labeling a protein with fluorophores using NHS ester derivitization. Methods Enzymol. 2014;536:87-94. |
| Additional Infomation | N-hydroxysuccinimde (NHS) ester-mediated derivitization involves the reaction of this amine-reactive group with the primary amines of a protein or a biomolecule. Using NHS chemistry allows one to conjugate various fluorescent probes, biotin, and cross-linkers to primary amines. For example, we use NHS ester chemistry to fluorescently label the amino terminus of a protein with the dye, 5-(and-6)-carboxyfluorescein, succinimidyl ester (5(6)-FAM, SE). [1] |
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.) |