Sephadex® G-100 (C09-1048-140)

Dextran gel is a kind of bead gel, which contains a lot of hydroxyl groups and swells easily in water and electrolyte solution. Hydrophilic matrix minimizes its non-specific adsorption, and achieves high recovery rate during biomolecule separation. G-type dextran gels have different crosslinking degrees, so their swelling degree and fractionation range are also different. The swelling degree of dextran gel is basically unaffected by the presence of salt and detergent.Instructions for use:  exist in the form of dry powder, which must be swelled before use. Excessive stirring should be avoided during swelling, because it may damage the filler. Do not use magnetic stirrer.I. Preparation of filler(1) expand the filler in excess deionized water or buffer solution at room temperature for 24 hours, or expand it with hot water for 1 hour (do not take a water bath! ）。Elution buffer should not contain high viscosity reagents. If there are floating objects on the upper layer during swelling, please remove them.(2) Balance the swelling filler, all buffers and other materials to the experimental operating temperature, and degas all buffers.Two-pack column(1) check whether all components of the chromatographic column, especially the filter screen, sealing ring and screw plug are tight, and whether the glass tube is clean and complete.(2) Wet the column and the bottom of the column with water or buffer and keep the liquid level for a short period of time, so that there is no bubble at the bottom.(3) Use a glass rod to guide the homogenate to be poured into the column at one time along the inner wall of the column, taking care not to generate bubbles. Open the liquid outlet of the column to make gelSettle freely in the column, and connect the top stigma of the column.(4) Open the peristaltic pump, and let the buffer flow through at a flow rate which is 1.33 times of the flow rate when in use, so as to stabilize the column bed (pay attention not to exceed the filling pressure)Material maximum pressure resistance).Three equilibriaBalance the chromatographic column at least 5-10 column volumes before loading until the baseline of the recorder becomes stable (pH value and conductance value of effluent are equal toPH value and conductance value of Buffer on the upper column).Four samplesSamples must be centrifuged or filtered (0.45um filter membrane).Generally, the loading amount of gel filtration is not more than 5% of the column bed volume, and we suggest that the initial loading should be controlled at 1-2% of the column bed volume, depending on the scoreFrom the situation can be adjusted; When desalting, the loading amount can reach 20% of the column bed volume, and the selection of column height is also related to the separation requirements. The higher the column, the higher the separationThe better the separation effect is, however, the gel column with too high column height will cause large back pressure, which should be avoided as much as possible. Difficult to separate substances must have certainColumn height and flow rate are controlled, and the ratio of height to diameter during desalination is 5: 1.Five elution methodsIt can be eluted with saline-free solution or buffer solution during column loading.Six cleaning in place (CIP)CIP is performed once after the gel is used ten times, in order to remove the precipitated and stubborn residual protein in the column bed. The method is to oxidize with 0.1 M hydrogen2 column volumes were washed with sodium, and then regenerated with at least 10 column volumes of equilibrium buffer.Add notes:(1) before loading, the sample must be filtered by membrane and pigment removed, otherwise impurities and pigment will be adsorbed on the filler and affect the fillerNormal use. All buffers need to be filtered with a 0.45um filter.(2) Avoid using strong acid and alkali with high concentration, and the concentration of acid and alkali should be lower than 0.1 mol. Alkali slows down the flow rate.(3) Different samples have different adsorption and elution methods, which can be carried out according to relevant literatures.Save notes:Untreated filler shall be sealed at room temperature. After using the filler, wash the salt thoroughly with pure water, and finally store it in 20% ethanol at 4℃Save. Specification: Medium MDL Number: MFCD00132235