Troubleshooting - High background
Immunohistochemistry (IHC) troubleshooting for no staining, high background and non-specific staining.
| Problems | Possible reasons | Solutions |
|---|---|---|
| High background | Antibody incubation time was too long | Decrease the antibody incubation time |
| Polyclonal primary antibody | Try a monoclonal primary antibody | |
| Serum blocking was absent or insufficient | Increase the blocking time | |
| High antibody concentration | Optimize the antibody concentration | |
| Presence of endogenous peroxidase | Block endogenous peroxidase activity properly | |
| Insufficient PBS washing | Wash with PBS for long enough and for enough times to eliminate fixatives |
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