HiTrap MabSelect SuRe英文说明书

GE Healthcare

Data File 11-0034-90 AC

Affinity chromatography

HiTrap MabSelect SuRe HiTrap MabSelect HiTrap MabSelect Xtra

HiTrap MabSelect SuRe (Superior Resistance), HiTrap

MabSelect , and HiTrap MabSelect Xtra (Fig 1) are members of the HiTrap family of prepacked columns for purification of monoclonal antibodies (MAbs).

Key performance characteristics of HiTrap MabSelect products include:

Increased binding capacity compared with other protein A media High purity in one step

Convenient, prepacked 1-ml and 5-ml columns Excellent for process development and screening of purification conditions Convenient operation with a syringe, a laboratory pump, or a chromatography system, such as ÄKTAdesign

The protein-A ligand used in all three MabSelect media is produced by validated fermentation and downstream processes. The entire production process is free of

components of mammalian origin. The resulting highly

purified ligand is immobilized to the agarose matrix through a chemically stable thio-ether linkage.

Fig 1. HiTrap MabSelect SuRe, HiTrap MabSelect, and HiTrap MabSelect Xtra 1 and 5 ml for purification of monoclonal antibodies with optimized binding capacities at high flow rates.

used for purification of different antibodies, as performing CIP will prevent cross-contamination between the different purifications. The enhanced alkali stability of MabSelect SuRe also improves purification economy; cleaning can be performed with cost-effective reagents such as 0.1 to 0.5 M sodium hydroxide.

The MabSelect SuRe ligand was developed by protein

engineering of one of the IgG-binding domains of protein A. Amino acids particularly sensitive to alkali were identified and substituted with more stable ones. The final construct is a tetramer of the engineered domain with a C-terminal cysteine, which enables single-point attachment to the matrix.The combination of low ligand leakage and high dynamic binding capacity together with the high-flow matrix makes MabSelect SuRe well suited for the purification of MAbs when scaling up. Characteristics of HiTrap MabSelect SuRe are listed in Table 1.

HiTrap MabSelect SuRe

MabSelect SuRe is composed of a rigid, high-flow agarose matrix and alkali-stabilized protein A-derived ligand that allows the usage of up to 0.5 M NaOH for cleaning-in-place (CIP). This ligand provides greater stability than conventional protein A-based media in the alkaline conditions used in CIP protocols. This is important when the same column is

imat work

DBC hIgG (mg/ml medium)

HiTrap MabSelect

MabSelect features a highly cross-linked agarose, produced using a manufacturing process that gives a very rigid matrix that is optimal for high-throughput affinity chromatography of IgG. The matrix of MabSelect allows at least five-times higher flow rates to be used in process scale compared with conventional cross-linked agarose of similar porosity. The epoxy-based coupling chemistry ensures low ligand leakage. The high capacity, low ligand leakage, and specially

developed base matrix make MabSelect ideal for purification 4035302520151051 ml/min0.4 ml/min

of monoclonal antibodies at process scale. See Table 2 for a summary of characteristics.

HiTrap MabSelect Xtra

MabSelect Xtra has been developed to meet the demands of ever-increasing levels of expression in monoclonal antibody feedstocks. MabSelect Xtra uses the same recombinant

protein A ligand as MabSelect, but has a smaller particle size and greater porosity, which ensures increased dynamic binding capacity. The medium provides a lower overall

production cost due to the possibility of processing concentrated feedstocks in fewer batches. The characteristics of HiTrap MabSelect Xtra are listed in Table 3.

Column characteristics

HiTrap columns are made of biocompatible polypropylene and they have porous top and bottom frits. Columns are delivered with a stopper on the inlet and a snap-off end on the outlet. Note that HiTrap columns cannot be opened or repacked.

Low ligand leakage

All three MabSelect media exhibit only a low level of ligand leakage during elution. Leakage is affected by chromatographic running conditions and the composition of the sample. For detailed information, please refer to the instructions for each media.

High dynamic binding capacities

The recombinant protein A in MabSelect media has been engineered to include a C-terminal cysteine. The coupling conditions are controlled to favor a thioether coupling providing single point attachment of the protein A. The oriented coupling also enhances the binding of IgG.

The dynamic binding capacity (DBC) of MabSelect media is affected by the flow rate, or residence time. The lower the flow rate (or the higher the residence time), the higher the DBC (Figs 2 and 3). A residence time of 2.4 min yields the specified binding capacities for the media, as stated in Tables 1 to 3. The dynamic binding capacity may decrease for columns with a bed height of 2.5 cm at the recommended flow rates (1 ml/min for 1-ml column or 5 ml/min for 5-ml column) because the residence time will be too

short to ensure optimal binding.

Data File 11-0034-90 AC

HiTrap MabSelect

HiTrap MabSelect Xtra

HiTrap MabSelect SuRe

Fig 2. Comparison of dynamic binding capacities (DBC) of HiTrap MabSelect, HiTrap MabSelect Xtra, and HiTrap MabSelect SuRe 1 ml columns at two different flow rates. Sample: 1 mg/ml human IgG (hlgG) Gammanorm (Octapharma); Binding buffer: 0.02 M phosphate, 0.15 M NaCl, pH 7.4; Elution buffer: 0.1 M sodium citrate, pH 3.0. Samples were run on an ÄKTAexplorer 10 system.

)

m40

uidem l35m/gm( G30gIh CBD2520151050

Residence time (min)

3.21.0

0.4

Flow (ml/min), 1 ml column16.75.0…… 此处隐藏：20640字，全部文档内容请下载后查看。喜欢就下载吧 ……