What are the causes of non-specific staining in tissue sections and how to resolve it?

1. Prolonged incubation or high antibody concentration may increase background staining; shorten primary/secondary antibody incubation and dilute the antibody. 2. Polyclonal primary antibodies tend to cause non-specific staining; monoclonal antibodies are recommended. 3. Endogenous peroxidase and biotin are abundant in liver, kidney and other blood-rich tissues; extend inactivation time and increase inactivating-agent concentration. 4. Non-specific components bind to antibodies; extend blocking with serum from the secondary-antibody host species and increase its concentration. 5. Excessively long DAB incubation or high DAB concentration increases background; reduce DAB concentration or shorten incubation. 6. Insufficient PBS washing leaves residual antibody and strengthens staining; wash thoroughly after primary antibody, secondary antibody and SP incubation. 7. Dried sections during staining often enhance non-specific staining; keep sections moist throughout. 8. Low pH of the antibody diluent or expired blocking serum.