The serum used to make this blocking solution is obtained from pools of healthy adult animals. The serum is heat inactivated at 56ºC for 60 minutes and dilpidated and filtered. The serum is tested with appropriate biotinylated antibody for cross-reactivity. Immuno protein blocking solutions is ready to use and do not contain azide or phosphate buffers.
- If tissues are used, deparafinize and hydrate tissue sections through xylene or other clearing agents and graded alcohols. (For frozen sections or cell smears; use unfixed, acetone fixed or appropriate fixative for the antigen in question; for cell smears it may be necessary to permealize the cell by detergent, please refer to antibody protocol).
- Wash 2-3 with distilled or deionized water.
- Incubate sections/cell smear in Endoblocker (#1) for 5-10 minutes at room temperature or 37°C. Note: If antigen retriever is required it can be applied after this stage.
- Wash slide with PBS or Tris saline (with 0.02-0.05% nonionic detergent, Triton X100, Tween 20 or NP-40) or washing buffer (Immuno Automation Buffer IBSC cat # AR-6561) 3-5X.
- Incubate sections/ cell smear in Immuno Protein blocking solution (diluted animal serum), for 10 minutes at RT or 37ºC or as suggested in your protocol.
- Wash slide with PBS 1X.
- Follow instructions for IHC.