Immunofluorescence (IF/ICC) Immunostain Protocol

Immunofluorescence Staining

1. Specimen was fixed with 4% formaldehyde in PBS for 15 minutes on ice.
2. Rinse three times in PBS for 5 minutes each.
3. Block specimen with 10% serum in PBS for 60 minutes at room temperature.
4. Incubate specimen with primary antibodies at 4℃ overnight.
5. Rinse three times in PBS for 5 minutes each.
6. Incubate specimen with fluorescence-labeled secondary antibody at 37℃ 1 hour.
7. Rinse three times in PBS for 5 minutes each.
8. Counterstained specimen with DAPI.
Note: If positive staining was localized to cytoplasm or nucleus, specimen was incubated with 0.3% Triton X-100 in PBS for 10 minutes after fixe