Application Description
Column packing
The Seplife LX-AG1×8 resin is shipped as wet resin. We recommend using pure water as the mobile phase for column packing. The recommended slurry concentration for column packing is 50-60%.
1) Gently mix the resins to form a homogeneous slurry; a resin volume of approx. 1.2 times more than the desired column packed volume should be used.
2) Before packing the column, ensure the slurry concentration in water is 50-60 %; pour the entre slurry quantity into the chromatography column.
3) Load the distributor and adjust the height, then start the pump and stabilize the column bed with 1.5 to 2 times the working flow rate.
4) Adjust the distribution plate height after the column bed is stabilized.
5) NOTE: the resin may shrink, or swell as much as 100%, depending on the (conversion) ionic form.
6) Efficiency and symmetry determinations are performed according to SOP and must meet predetermined criteria.
Equilibration
After packing the column, equilibrate with the mobile phase first, with 5-10 column volumes, and control the flow rate at 120-300cm/h until the conductivity and pH of the flow-through remain unchanged before feeding the sample. For peptide conversion salt format, 0-50% acetonitrile is recommended as the mobile phase.
Sample feeding
The solid sample can be prepared by dissolving in the equilibrium solution. The feed amount is calculated according to the
capacity of the resin and content of the feed solution. Before loading, ensure that the sample buffer should be as consistent as possible with the equilibration solution. For peptide conversion salt format, 0-50% acetonitrile is recommended.
Elution
After loading the sample, continue rinsing with equilibration buffer until the baseline is stable. According to the actual situation, elute the samples adsorbed on the resin sequentially by increasing the salt concentration or changing the pH of the mobile phase. For peptide conversion salt format, 0-50% acetonitrile is recommended as the mobile phase.
Regeneration and CIP
Regular Cleaning-In-Place (CIP) can prevent column fouling and help to maintain the capacity and separation effect of the
chromatographic media. Specific CIP methods and the frequency of CIP need to be designed for each process according to the type of contamination. The recommended regeneration and CIP method is as follows: Rinse the column up flow with 5 CV of 1.0 NaCl, 0.1 M NaOH in 20% acetonitrile.
Resin conversion
To convert a resin to an ionic form with a higher selectivity, wash the resin with 2-5 bed volumes of a 1 M solution of the
desired counterion. For conversion to an ionic form with a lower relative selectivity for the resin, the necessary volume of counterion solution will depend on the difference in selectivity. Common techniques for converting ion exchange resins from one ionic form to another.