Description
Detailed procedure
After cell lysis His-tagged proteins are magnetically labeled with µMACS Anti-His MicroBeads. The sample is loaded onto a MACS Column placed in the magnetic field of a µMACS or thermoMACS Separator in which the magnetically labeled His-tagged and associated proteins are retained during the washing steps. After elution with denaturing elution buffer, the precipitated proteins can be analyzed by SDS-PAGE and Western blot. Alternatively, native proteins can be eluted by pH-shift using triethylamine pH 11.8 or by removing the column from the magnetic field and applying a suitable native elution buffer.
Applications
The µMACS and MultiMACS His Isolation Kits facilitate the isolation of His fusion proteins from different sources. The super-paramagnetic µMACS MicroBeads included in the kits are conjugated to an anti-His monoclonal antibody, enabling fast and effective magnetic labeling of His-tagged fusion proteins. The protocol allows for protein isolation within just 1.5 hours. No centrifugation or buffer removal steps are required.
Downstream applications
The (co-)immunoprecipitated proteins can be analyzed by SDS-polyacrylamide gel electrophoresis, Western blot analysis, or mass spectrometry1. Enzymatic on-column assays can also be performed using the thermoMACS Separator.
Overview
The µMACS and MultiMACS His Isolation Kits facilitate the isolation of His fusion proteins from different sources. The super-paramagnetic µMACS MicroBeads included in the kits are conjugated to an anti-His monoclonal antibody, enabling fast and effective magnetic labeling of His-tagged fusion proteins. The protocol allows for protein isolation within just 1.5 hours. No centrifugation or buffer removal steps are required.
