Tandem Affinity Purification (TAP) Protocol (Page 5)
- Check by PCR the presence of the TAP tag on your protein of interest.
- Do a small prep to make sure your protein is in the soluble fraction.
- Check health of cells with the TAP tag by doing a growth curve. A healthy haploid strain doubles between 80-90 minutes.
- Wear gloves to protect your samples from keratin, which can show up in mass spec.
- Test activity of TEV protease to ensure good cleavage off the IgG column. Commercial TEV preparations do not always work well.
- Make sure samples are kept cold throughout the purification, especially during lysis.