Section8:Reagents

From Assay Guidance Wiki

Contents 1 Plates 2 Coating 3 Fixatives 4 Antibodies 5 Buffers 6 Substrates

Plates

• White plates for chemiluminescent readout
• Black plates for fluorescent readout
• Clear plates if reading colorimetric

Coating

• Poly-D-lysine
• Collagen
• Gelatin
• Fibronectin

Fixatives

• 3.7% Formaldehyde (Sigma catalog #F1635 37% solution) in PBS
• Commercially available non toxic fixative reagents
• Methanol

Antibodies

• Primary antibody specific for protein of interest
• Secondary antibody:
  • For chemiluminescence: anti-rabbit or anti-mouse IgG HRP conjugate
  • For fluorescence detection using the high content imaging technology use: anti- mouse or anti-rabbit IgG Alexa 488
  • For fluorescence detection using the the infrared use the reagents labeled with IR tags

Buffers

• Phosphate buffered saline
• Wash buffer: PBST (PBS with 0.1% Triton X-100)
• Antibody dilution and blocking buffer: PBS with 0.5% BSA and 0.1% Triton X-100

Substrates

• Colorimetric: TMB
• Chemiluminescent: Commercially available chemiluminescent reagents