I am trying to purify a cell lysate using a 1 mL His-trap column but the viscosity of the lysate is clogging up the column. What are some solutions to this?
Treat the lysate with DNase + MgCl2 to reduce the viscosity caused by DNA. Centrifuge the lysate before applying it to the column to remove any particulates.
LYSATES CAN BE FILTERED THROUGH STERILE 0.22 MICRON OR ATLEAST 0.45 MICRON STERILE FILTERS BEFORE INJECTING INTO COLUMN , IF THE INJECTABILITY IS LESS THEN CAN BE DILUTED WITH RESUSPENSION BUFFER AND CAN BE FILTERD AND INJECTED INTO THE COLUMN , THIS CAN AVOID CLOGGING
I agree with the suggestion from Adam B Shapiro. As well as using DNAse I, centrifuge the lysate at 10,000 g or greater for at least 10 minutes (the longer and faster the better at sedimenting aggregates/insoluble material that can clog the column) and put it through a 0.22 micron syringe filter. If it is still viscous, you might need to increase the volume of your lysis buffer at the lysis stage or dilute the lysate a bit before applying to the column.
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