Cell Lysate ELISA - Find protocol for a cell-lysate ELISA (Jan/24/2002 )
I'm interested in doing a sandwich ELISA to a cell lysate. Does anyone have a protocol describing this? Most importantly I'm looking for the types of buffers used in such an assay.
I guess not many people are working on this issue. I too tried to create a cell lysate ELISA with little success. Was hoping to pick up my his-tagged heterologously induced protein from the soup using and anti tetra-his antibody, but don't get a signal above background. Anyone have any luck with this approach?
I assume you did an extensive dilution series? Sometimes the lysate has interfering compounds.
As for the His-tagging expt, does your protein purify happily by binding to NTA beads? I know a lot of people have had problems with proteins not binding to their beads. You might need to play with your conditions. If your can get conditions where the protein purifies on beads, they might be good starting conditions for the ELISA.