Dextrin Beads 6FF By Lablead biotech,

Product Specification

Product Description

Dextrin Beads 6FF CAT：M***0

Storage：**8 °C
Price: $**0 for *0ml; $**0 for **0ml (negotiable)
Product Introduction
Dextrin Beads 6FF is an affinity chromatography medium for the purification of maltose-binding protein (MBP) -tagged proteins, and the specific properties are shown in Table 1. MBP promotes the proper folding of connexins and increases the solubility of fusion proteins that are overexpressed in bacteria, especially eukaryotic proteins. Dextrin Beads 6FF could purify the MBP fusion protein in one step, and the bound fusion protein could be gently eluted with *0 mM maltose, protecting the activity of the tag protein. If you want to remove the MBP fusion part, you can use site-specific protease excision.
Table 1 Product Performance of Dextrin Beads 6FF

Item	Property
ligand	Highly crosslinked 6% agarose microspheres
Matrix	dextrin
capacity	>0 mg MBPprotein(0 kDa) /ml resin
Particle	*****5 pm
maximum pressure	0.3 MPa, 3 bar
pH stability range	***2
Storage buffer	1xPBS containing *0% ethanol

Process of purification
1 The buffer
Before using the water and buffer suggest using 0.*2 um and 0.*5 um filter membrane filtration.
Equilibration/washing buffer: *0 mM Tris-HCI, **0 mM NaCl, 1 mM EDTA, pH7.4;
Elution buffer: *0 mM Tris HCl, EDTA, 1 mM to *0 mM maltose, pH7.4;
Note: 1mM DTT or *0mM β-hydrophobic ethanol can be added to equilibration and eluate
2 Sample preparation
Samples are recommended to be centrifuged or filtered with a 0.*2um or 0.*5um filter membrane before loading to reduce impurities, improve protein purification efficiency and prevent blocking of the column.

3 Dextrin Beads were loaded with 6FF
3.1 Loading of gravity column
1) Take the gravity chromatography column of appropriate specifications, load the lower gasket, add an appropriate amount of pure water to wash the column tube and gasket, and close the lower outlet.
2) the Dextrin Beads 6 ff mix, use spear absorb right amount in the slurry to join to the gravity column (medium actual volume accounts for about half of the suspension), open the outlet flow under dry protection fluid.
3) Add appropriate amount of pure water to rinse the medium, and close the lower outlet after the liquid in the column is dried by gravity flow.
4) Load the upper gasket after moistening and washing, ensure that there is no gap between the gasket and the filler, and keep it level.
5) The loaded gravity column can be directly balanced by adding the balance solution, and the protection solution can be added when not in use temporarily and stored at **8℃.
3.2 medium pressure chromatography column packing
Dextrin Beads 6 ff purification is widely used in industry, therefore, involves a variety of medium pressure chromatography chromatography column fill, fill in the method of chromatography column is described below. Before column loading, the column base area was calculated according to the diameter of the chromatographic column, and the required medium volume was calculated according to the required column loading height. The formula is as follows:
(V: V = 1.*5 PI r2h the required volume of medium ml; 1.*5: the compression coefficient; R: column pipe radius cm; H: loading height cm)
Note: the suspension liquid product should be two times the size of medium, because of suspension medium volume half of the total volume, the other half to protect fluid.
1) Wash the bottom sieve plate and joint of the chromatography column with deionized water to ensure that there are no bubbles on the sieve plate at the bottom of the column, close the outlet of the bottom of the column, and leave **2 cm of deionized water at the bottom of the column.
2) medium is suspended, carefully serous continuously in chromatography column. The formation of bubbles was reduced by pouring slurry along the column wall with a glass rod.
3) If a reservoir is used, fill the chromatographic column and reservoir with water immediately, place the injection dispenser on the slurry surface and connect it to the pump to avoid air bubbles in the dispenser or the injection tube.
4) Open the bottom outlet of the chromatography column and open the pump so that it can proceed at the set flow rate. Buffer should initially be allowed to flow slowly through the column, and then slowly increased to the final flow rate, so as to avoid the impact of hydraulic pressure on the resulting column bed and to avoid the formation of non-uniform column bed. If the recommended pressure or flow rate is not reached, the maximum flow rate of the pump used can be used, and a good filling effect can also be obtained. (Note: In the subsequent chromatographic procedure, do not exceed *5% of the maximum column loading flow rate.) When the column bed is highly stabilized, at least 3 times the column bed volume of deionized water is added at the final column loading flow rate. Mark the height of the column bed.
5) Turn off the pump and close the column outlet.
6) If a reservoir is used, remove the reservoir and place the dispenser in the chromatographic column.
7) Push the dispenser towards the post to the height of the marked post bed. Allow the column loading fluid to enter the dispenser and lock the dispenser joint.
8) Connect the loaded chromatographic column to the pump or chromatographic system and begin equilibration. The allocator can be readjured if needed.

4. Sample purification process
4.1 Purification by incubation
1) According to the amount of purified sample, an appropriate amount of Dextrin Beads 6FF was added to a centrifuge tube, centrifuged at ***0 RPM for 1 min, and the supernatant was absorbed and discarded. It can also be added to the gravity column to dry the protection solution.
2) Add 5 times the volume of medium balance solution to the centrifuge tube to clean the medium, centrifuge at ***0rpm for 1 min, and discard the supernatant; Column, such as using the gravity directly cleaning in the column of gravity, gravity flow directly dried liquid balance; Repeat more than twice.
3) to join the samples, enclosed centrifugal pipe or gravity column, 4 ° C oscillation **4 h incubation or *7 ° C incubation for *0 min - 2 h.
4) after the incubation, ***0 RPM centrifugal 1 min, abandon supernatant, or filter collection medium, supernatant retained as flow wear, used in electrophoresis identification.
5) Wash the medium with 5 times the volume of the medium, centrifuge at ***0 rpm for 1 min or filter with a gravity column tube to remove the supernatant (be careful not to absorb the medium), repeat **5 times, and it is recommended to replace a new centrifuge tube in the middle.
6) Add **5 times the column volume of eluate for elution, incubate at room temperature for ****5 min, centrifuged at ***0 rpm for 1 min or collect the eluate by gravity column tube, which can be repeated for **3 times.
4.2 Gravity column purification
1) The loaded Dextrin Beads 6FF gravity column was equilibrated with 5 times the column volume equilibrating solution so that the filler was under the same buffer system as the target protein and repeated **3 times.
2) Add the sample to the well-balanced gravity column, and the sample retention time is at least 2 min to ensure that the sample and the medium are fully in contact, and the effluent is collected with repeated sample increase combining efficiency.
3) Wash with ****5 times the column volume, remove the non-specific adsorbed miscellaneous proteins, and collect the miscellaneous solution.
4) The eluate was eluted with ***0 times the column volume, collected in sections, one tube for each column volume, and detected separately, so that all bound target proteins could be eluted, and high purity and concentration of proteins could be obtained.

4.3 Purification by medium pressure chromatography
After loading, Dextrin Beads 6FF can be used with a variety of conventional medium and low pressure chromatography systems.
1) Fill the pump pipe with deionized water. Remove the top plug, connect the column to the chromatographic system, open the lower outlet, attach the preloaded column to the chromatographic system, and tighten it.
2) The storage buffer was flushed out with **5 times the column volume of deionized water.
3) use at least five times the column bed volume balance liquid equilibrium chromatographic column.
4) Sample loading by pump or sample ring.
Note: The increased viscosity of the sample causes a large backpressure on the chromatographic column even if the loading volume is small. The loading volume should not exceed the binding capacity of the column. The large sample volume may also cause a large backpressure, making the injector more difficult to use.
5) with the washing of mixed liquid or pillars, until the ultraviolet absorption reaches a stable baseline (typically at least ****5 column volumes).
6) The eluate was eluted using ***0 times the column volume and the eluate, which was the target protein fraction, was collected.
The above steps after the elution, flush with balance fluid column volume 3 times, 5 times and then rinse with water column volume, reoccupy protective liquid washing 2 column volume, and then puts medium **8 ° C to save.

5. SDS-PAGE detection
The samples obtained using the purified product (including effluent fraction, washed fraction and eluted fraction) and the original sample were tested by SDS-PAGE.

6. Packing to clean
Dextrin Beads 6FF purified products can be reused without regeneration, but the increase of non-specific binding proteins and the aggregation of proteins often cause the flow rate and binding load to decrease. At this time, the packing can be cleaned as described below.
1) 3 times the column volume of deionized water;
2) three times column volume of 0.1% SDS or 0.1 M NaOH solution;
3) Three times the column volume of deionized water and *0% ethanol were stored at **8℃.

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Product Group :	Protein Biology