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Product Introduction
Q-agarose gel HP is a strong anion exchange medium formed by bonding quaternary ammonium groups on agarose microspheres. It has the characteristics of high resolution, high load capacity, high recovery rate and strong regeneration ability. Good chemical stability, can be used in sodium hydroxide cleaning. The hydrophilic properties of the matrix ensure that non-specific adsorption of cell-derived impurities is reduced during elution. It can be used for ion exchange preparation of protein, nucleic acid and polypeptide.
Product parameters
Product Name: Q-Sepharose HP (Q-Sepharose High Performance)
Item number: S9200
Substrate: 6% crosslinked agarose gel
Ligand: Quaternary ammonium
Exchange group: -N +(CH3)3
Ionic capacity: 0.14-0.20 mmolCl-/mL
Particle size: 34μm
Protein load: 10mg lgG, 24mg HAS/mL
Working pH: 1~14(short time, in-place cleaning); 2~12(Long time)
Chemical stability: Stable in the following solution -1mol /L NaOH; 8mol/L urea; 6mol/L guanidine hydrochloride; 30% isopropyl alcohol; 70% ethanol; 30%SDS, 30% acetonitrile;
How to use
1. Install the column
(1) Bring all reagents and fillers to room temperature. Prepare the initial buffer (balance) and buffer.
(2) Take the amount of gel required according to the size of the column, wash off 20% ethanol, drain it, and prepare a homogenate with the initial buffer (according to the gel: buffer =3:1 ratio).
(3) Wet the inside of the column and the bottom of the column with water or buffer and keep a small section of liquid (the liquid level is slightly higher than the filter membrane), be sure to make the bottom end free of bubbles.
(4) Use a glass rod to guide the homogenate along the inner wall of the column into the column at one time, taking care not to use bubbles. Open the liquid outlet of the column, make the gel settle freely in the column, and connect the top of the column.
(5) Open the peristaltic pump, so that the buffer flow rate of 1.33 times the flow rate of use, so that the column bed is stable. Balance the column with a buffer of 2-3 times the column volume.
Step 2 Balance
Let the equilibrium buffer flow through the column at a constant flow rate until the effluent conductance and pH are unchanged. The equilibrium buffer is generally a buffer with a high salt concentration, such as Tris, PBS, etc.
3. Sample delivery
(1) The sample is prepared with equilibrium liquid, and the turbid sample should be centrifuged and filtered on the column;
(2) In general, the target product is combined on the column, the impurities are washed with a balanced lotion, and then an eluent is selected to wash the target product.
(3) The degree of adsorption of the sample components depends on the charged properties of the sample, the ionic strength of the mobile phase and the pH value. The smaller the salt concentration, the more strongly the medium adsorbed the components.
Step 4 Elution
Q The agarose gel medium can be eluted by increasing the salt concentration or decreasing the pH value, and the method of increasing the salt concentration is commonly used.
Step 5 Regenerate
Generally washed with a buffer of high salt concentration (containing 1 to 2mol/L NaCl) or reduced pH to wash more than 10 times the column volume, and then washed with a balance solution of binding proteins to balance, can be used again.
If deactivated proteins or lipids cannot be washed off during regeneration, they can be removed by in-place cleaning (CIP).
6. Clean while in position
(1) Proteins bound by ionic bonds can be removed with 2M NaCl.
(2) For precipitated proteins, proteins or lipids bound by hydrophobic action, 1M NaOH can be removed.
(3) For strongly hydrophobic binding proteins, lipids, etc., can be cleaned with 70% ethanol or 30% isopropyl alcohol of 4-10 times the column volume, but pay attention to the gradual increase in the concentration of organic solvents in a gradient manner, otherwise it is easy to produce bubbles.
After cleaning, balance the column with at least 3 times the column volume buffer.
7. Remove from heat source
Clean the column with 0.5M of sodium hydroxide for 5-6 hours or 0.1M of sodium hydroxide for 24 hours. Or remove with the following steps:
(1) Twice the column volume of 70% ethanol;
(2) Twice the column volume 50Mm Tris-HcL Ph7.5
(3) 1 column volume 4M urea
(4) Tris buffer 3 times the column volume +0.1M NaCl;
The above buffers are prepared in double steaming water without heat source.
Step 8 Disinfect
Wash 8-10 times the column volume with 0.5-1.0M NaOH at room temperature to balance the column with the initial buffer.
Precautions
(1) Sealed storage in 4℃-28℃(storage solution is 20% ethanol), ventilated, dry place, can not be frozen. The used columns are stored at 4℃(20% ethanol).
(2) When installing the column, using and preserving the column, it is necessary to avoid the column drying and air bubbles entering.
Note：Product information may be optimized and upgraded. Please refer to the actual label information for accuracy.
