pH Gradient Buffer Solutions
Recombinant monoclonal antibodies (MAbs) can be highly heterogeneous due to modifications such as sialylation, deamidation and C-terminal lysine truncation. As biotherapeutic drugs are becoming more popular, chemists in biopharmaceutical laboratories are coming under increasing pressure to adopt a fast, generic, and robust approach to clone screening, method development, and subsequent method transfer to QA/QC.
Traditionally cation-exchange chromatography using salt gradients has been successfully used to characterize MAb charge variants. However, additional effort is often required to tailor the salt gradient method for individual charge variants. Thermo Scientific™ pH buffer solutions and kits can be used to generate highly reproducible, linear pH gradients using cation-exchange chromatography. This generic, LC-based platform approach saves time in method development and facilitates method transfer to QA/QC for a wide range of MAb charge variants. Unlike traditional salt gradients, it is possible to predict the pI and the expected retention of the charge variants and use a narrow pH range to get a higher resolution separation.
The building blocks of the pH gradient platform are two multicomponent zwitterionic buffer concentrates, prepared using a patent-pending formulation. Buffer A is titrated to pH 5.6 and Buffer B is titrated to pH 10.2. In this pH range, each buffer species is either neutral or negatively charged. Therefore, they will not be retained by the cation-exchange column stationary phase and will serve as good buffers for the mobile phase and the stationary phase. All that is required to generate a pH gradient is a 1:10 dilution of the pH buffer concentrates—then you’re ready to go! A linear pH gradient from pH 5.6 to 10.2 can simply be delivered by running a pump gradient from 100% eluent A to 100%eluent B.
Thermo Scientific pH buffer concentrates used in the pH gradient platform can be purchased individually or as a pair, in quantities of 125 mL or 250 mL. For added convenience, the 125 mL buffers can also be bundled with columns in a number of specifically preconfigured kits.
The CX-1 pH gradient starter kit contains 125 mL each of buffers A and B, plus a MAbPac SCX-10, 10 μm, 4 × 250 mm column.
The CX-1 pH gradient high throughput kit contains 125 mL each of buffers A and B, plus a MAbPac SCX-10, 5 μm, 4 × 50 mm column.
The CX-1 pH gradient high resolution kit contains 125 mL each of buffers A and B, plus a MAbPac SCX-10, 5 μm, 4 × 250 mm column.
For the ultimate flexibility, the preconfigured kits are also available as platforms, including the pH Designer Software. The options are listed in the table below:
pH Buffer Concentrates |
---|
CX-1 pH Gradient Buffer A (pH 5.6), 125 mL | 083273 |
CX-1 pH Gradient Buffer B (pH 10.2), 125 mL | 083275 |
CX-1 pH Gradient Buffer Kit (pH 5.6 to 10.2), 125 mL | 083274 |
CX-1 pH Gradient Buffer A (pH 5.6), 250 mL | 085346 |
CX-1 pH Gradient Buffer B (pH 10.2), 250 mL | 085348 |
CX-1 pH Gradient Buffer Kit (pH 5.6 to 10.2), 250 mL | 085349 |
CX-1 pH Gradient Starter Kit (pH 5.6 to 10.2), 125mL | 083381 |
CX-1 pH Gradient High Throughput Kit (pH 5.6 to 10.2), 125 mL | 083378 |
CX-1 pH Gradient High Resolution Kit (pH 5.6 to 10.2), 125 mL | 083272 |
CX-1 pH Gradient Starter Platform (pH 5.6 to 10.2), 125 mL | 083380 |
CX-1 pH Gradient High Throughput Platform (pH 5.6 to 10.2), 125 mL | 083376 |
CX-1 pH Gradient High Resolution Platform (pH 5.6 to 10.2), 125 mL | 083270 |
pH Designer Software | 085022 |
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