C. elegans Immunoprecipitation

Contributed by Ian Chin-Sang, Queens University, ON, Canada

Immunoprecipitation protocol for use with C. elegans worm lysate

  • Preparation of worm lysate:
  • Add 2 ml of PLC buffer to 0.25 ml of worm pellet.
  • Break up the worms by sonicating.
  • Spin 10 minutes @ 13000 g in microfuge, save supernatant.
  • Quantitate protein in lysate using Pierce BCA kit.
  • Wash 100 ml (actually 10 ml of beads since it comes as a 10% suspension) of Protein A Sepharose in PLC buffer. Spin 3 minutes @ 13000 g in microfuge.
  • Immunoprecipitate using 1 mg of lysate protein (may have to use more depending on the protein you’re looking at) mixed with 5 mg of antibody and 10 ml of the washed Protein A Sepharose. Total volume of the mixture should be 1 ml.
  • Incubate on a rocker platform at 4ºC for 1 hour.
  • Wash 3x with 1 ml of PLC buffer.
  • Add 50 ml 1x sample buffer.
  • Boil 5 min.
  • Load 20 ml on an SDS PAGE gel.

This method is based, with permission, on an original protocol available here.