Protocols - Immunohistochemistry: Mouse Anti-Human MCP-1
Immunohistochemistry protocol for Mouse Anti-Human MCP-1*:
The following protocol used formalin-fixed paraffin-embedded human tonsil tissue (supplied by the Alfred Hospital, Australia).
- Deparaffinize and rehydrate the tissue section.
- Perform heat-induced antigen retrieval with 1mM EDTA pH 8.0 at 123˚C for 3 minutes in a Pascal pressure chamber (Dako). Wash the slide twice for three minutes each (50mM Tris-HCl pH 8.0, 150mM NaCl in distilled water).
- Incubate the tissue section with 1% BSA/PBS block.
- Incubate the tissue section overnight at 4˚C with Mouse Anti-Human MCP-1 at 10.0 μg/mL. Wash the slide twice for three minutes each.
- Incubate the tissue section with the Envision+ HRP polymer kit (Dako) according to the manufacturer’s protocol. Wash the slide twice for three minutes each.
- Incubate the tissue section with the DAB chromogen.
- Counterstain the tissue section with hematoxylin.
- Coverslip the sections with DPX mounting medium and image sections with an Olympus Provis light microscope (Monash Micro Imaging, Monash University, Australia).
* Protocol and staining provided by Dr. Lauren Binge, Laboratory of Prof. Charles Mackay, Monash Univeristy, Australia.