Guidelines (This is our suggested protocol; it should be adjusted based on your unique requirements as it just offers as a guideline). 1. Swelling: It is necessary to swell the dry rubber particles initially. Stem excess deionized water and let it swell for three hours at room temperature or for one hour in a water bath at ninety degrees Celsius. (To prevent fracturing the microspheres during the swelling phase, magnetic stirring is prohibited.) 2. Column packing: (1) Based on the characteristics of the samples to be separated, prepare the elution and initial buffers (equilibrium solution and initial buffer). It is usually advised to utilize a single buffer with a low salt concentration for sample loading, equilibration, and elution in gel chromatography for desalting proteins, such as proteins. For instance, 50 mM Tris-hydrochloric acid or PBS buffer close to neutrality; if peptide separation is the goal, 0.15 M NaCl can be suitably added to prevent target-gel contact; if buffer exchange is the goal, choose the target buffer as required. (2) After draining the gel, make a homogenate with the first buffer (3:1), then degas. (3) Fix the column vertically, moisten the lower end with buffer or water, and maintain the liquid level for a while. (4) Using a glass rod as a guide, pour the homogenate into the column one time around the inner wall, letting the gel settle freely. Then, cover the column with water and let it settle for the night. (5) Attach the movable column head at the top of the column, activate the peristaltic pump, allow the buffer to flow through five times the column volume using the operating flow rate, and then add 1.5 times the operating flow rate to allow the buffer to flow through five times the column volume, adjusting the adapter head to the closest setting. Lastly, use two to three column volumes of buffer to equilibrate the column. Note: To guarantee the gel's homogeneity, air bubbles cannot be added during any step of the process. 3. Equilibration: Using the equilibration buffer, equate the column at the operating flow rate. Then, watch how the detector changes until the conductivity and pH levels don't change. 4. To load a sample, turn on the switching valve. The type of sample and chromatography medium quantity are taken into consideration while choosing the sample load. In general, the sample load doesn't go above 25% of the volume of the column bed. The samples underwent pretreatment by filtering through a 0.45 μm microporous membrane after being prepared with an equilibration solution. 5. Elution: After peaking, switch to an elution buffer to achieve consistent elution to baseline equilibrium. 6. Regeneration: Rinse with deionized water for three to five times the column volume after washing with 0.2 mol/L NaOH or non-ionic detergent for two times the column volume. 7. Storage: After using, swap out the mobile phase for 20% ethanol and keep the mixture in a 4°C refrigerator. 8. Maintaining column heights of various particle sizes at 30 cm and above for coarse particles, 20 cm and above for medium particles, 10 cm and above for fine particles, and 5 cm and above for ultra-fine particles is advised in order to achieve an improved desalination effect. The smaller the particle size, the slower the flow rate and the higher the resolution. 9. The experiment should be used to determine how much sample is added. The ideal sample load is between 10% and 20% of the column bed volume, with a maximum load of 30% not to be exceeded.

[1]. Saul A, et al. A rapid method of concentrating proteins in small volumes with high recovery using Sephadex G-25. Anal Biochem. 1984 May 1;138(2):451-3.

0

9041-35-4

Typically exists as solids (or liquids in special cases) at room temperature

O1[C@]([H])([C@@]([H])([C@@]([H])([C@@]([H])([C@@]1([H])C([H])([H])O[H])O[H])O[H])O[H])O[H].O([H])C([H])(C([H])([H])O[H])C([H])([H])O[H]

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

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

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

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.

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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).

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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)

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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).

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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

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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.

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