Anti-HA-Fluorescein Troubleshooting

Product No. 11666878001

Troubleshooting

Signal week or missing; too few cells stained

  • Protein is not expressed in cells. → Prepare a cell extract and use western blot analysis to verify that tagged protein is expressed.
  • Too few cells expressing protein → Use more cells in sample. Try a different transient transfection procedure or use a stably transfected cell line.
  • Poor cell permeability → Increase time of incubation with PBS/Triton.
  • Sample fixation before staining destroys epitope → Use a different sample fixation method.
  • Tag-specific antibody too dilute → Run a titration curve (identical samples, varying amounts of tag-specific antibody, optimal amounts of secondary antibody) to determine the optimal dilution of tag-specific antibody.

High background

  • Antibody too concentrated → Run a titration curve (identical samples, varying amounts of tag-specific antibody, optimal amounts of secondary antibody) to determine the optimal dilution of tag-specific antibody.
  • BSA in Blocking Solution contains IgG → Use only high purity (IgG-free) BSA.

Materials

     
Related Links