Sephadex G-25 medium

Description: Separation reagent, chromatographic medium. Separation range: 1000-5000

Chemical and Physical properties :

Glucan gel is a beaded gel that contains a large number of hydroxyl groups, which swell easily in water and electrolyte solutions. G-type dextran gels have a variety of crosslinking degrees, so their swelling and fractionation ranges are also different. The swelling degree of dextran gel is basically unaffected by the presence of salt and detergent.

Dextran gels come in different particle sizes. Ultrafine grade dextran gels are used for column and thin-layer chromatography requiring extremely high resolution. Coarse and intermediate gels are used in preparative chromatography processes to achieve higher flow rates at lower pressures. In addition, coarse grades can also be used for batch processes.

Chemical stability :

Dextran gel is insoluble in all solvents (unless it is chemically degraded). It is stable in water, salt solution, organic solvent, base and weakly acidic solution, and the glycosidic bonds of the gel skeleton are hydrolyzed in strong acid. Prolonged exposure to oxidants will destroy the gel and should therefore be avoided.

Physical stability :

Dextran gum does not melt and can be sterilized at wet, neutral PH or in an autoclave at 120℃for 30 minutes without affecting its chromatographic properties. The dry gel will begin to caramelize when heated above 120℃. The mechanical strength of dextran gels depends on the degree of crosslinking.

Usage :

1. Glucan gel pretreatment:

At room temperature, soak dextran gel powder in distilled water for at least 24 hours, stirring constantly to ensure gel swelling, or soak in hot water (boiling is not recommended) for at least 1 hour until fully swollen and the gel volume does not change.

Note: Before swelling, add distilled water and stir it to make it settle naturally. After settling, if there are more gel fragments floating in the upper solution, it is necessary to rinse them repeatedly to remove them, so as to prevent blocking of the gel column during chromatography and affecting the flow rate.

2. Column loading:

Place the swelling gel into the column at one time according to the requirements of column loading, pay attention to keep the wet state of column loading, and avoid bubbles or faults in the column; The column should be uniform, and the column can be pressed down in the operation of gel tolerance. The gel column can be installed to test the column effect and test whether the filter column is qualified.

The gel chromatographic separation degree depends on the column height, in order to separate different components, the gel column bed must have a suitable height: in the fractional separation, the chromatographic column is generally about 100cm long, and the ratio of column height to diameter is 20:1-100:1. When the column height is too high, the gel extrusion deformation and blockage will cause large back pressure, which should be avoided as much as possible. For group separation (such as desalting), a short column is used, the column height is controlled below 40-50cm, and the ratio of column height to diameter is about 5:1-10:1.

3. Balance:

Balance the chromatographic column for at least 3-5 column volumes with the equilibrium solution before loading until the baseline of the recorder becomes stable (e.g., the PH of the effluent is equal to the PH of the Buffer of the upper column);

4. Sample delivery:

The loading amount is generally 5% of the column bed volume in the stage separation, and we recommend that the initial loading control be 1-2% of the bed volume, which can be adjusted according to the separation situation. During desalting, the loading amount can reach 20% of the column bed volume.

If the sample solution has impurities or precipitation, it should be filtered or centrifuged, and the viscosity of the sample should not be too high, otherwise the separation effect will be affected.

5. Elution method:

It can be eluted with no salt water or with the buffer solution in the column. The separation and purification can also be achieved by adding NaCl or salt gradient elution into the upper column buffer.

6. In-place Cleaning (CIP) :

The gel is applied ten times for a CIP to remove the precipitated and persistent proteins in the column bed. The method was to reverse clean four column volumes with 1M sodium hydroxide at 40cm/h, and then regenerate with at least three column volumes of balanced buffer.

7. Save:

The gel column is not used for the time being, it can be recycled, the general method is to rinse the gel with water and drain it, can be added 0.02% sodium azide preservative or soaked in 20% ethanol to control microbial growth.

Note：Product information may be optimized and upgraded. Please refer to the actual label information for accuracy.

Remark：These protocols are for reference only. Solarbio does not independently validate these methods.

Note：

1. The products are all for scientific research use only. Do not use it for medical, clinical diagnosis or treatment, food and cosmetics, etc. Do not store them in ordinary residential areas.

2. For your safety and health, please wear laboratory clothes, disposable gloves and masks.

3. The experimental results may be affected by many factors, after-sale service is limited to the product itself and does not involve other compensation.