Table 1.
Proteins, source and dilution of antibodies, staining procedures and patterns and interpretation of results
| Protein | Antibody [clone, source] | Ab dilution | Pretreatment/EU | IT | IHC Staining Detection System/Chromogen | Scoring System | Cut-off (%) | Staining Pattern | Reference |
|---|---|---|---|---|---|---|---|---|---|
| Ki67 | MIB1 (1) | 1:70 | 15'/EU2 | 20' | Polymer HRP/DAB | SQ | ≥ 14%* | Nuclear | [28] |
| HER2 | PL (1) | 1:200 | 25'/EU1 | 30' | Polymer HRP/DAB | 0-3 | > 30%# | Membranous | [23] |
| TopoIIa | KiS1 (1) | 1:200 | 15'/EU2 | 30' | EnVision/DAB | SQ | > 5%** | Nuclear | [27] |
| ER | 6F11 (2) | 1:70 | 20'/EU1 | 20' | Polymer HRP/DAB | H-Score | ≥ 1% | Nuclear | [29] |
| PgR | 1A6 (2) | 1:70 | 20'/EU1 | 20' | Polymer HRP/DAB | H-Score | ≥ 1% | Nuclear | [29] |
Antigen Retrieval was done on a hot plate at 98°C.
DAB: 3,3'-Diaminobenzidine; EU: Epitope Unmasking; EU1: Citric acid, pH 6.0; EU2: Ethylenediaminetetraacetate, pH 8.8; HRP: Horseradish Peroxidase; IT: Incubation time; SQ: Semiquantitative.
(1): Dako, Glostrup, Denmark; (2): Leica Biosystems, Newcastle Upon Tyne, UK.
*: Proliferation Index: Low if < 14% and High if ≥14% positive nuclei were observed; #: score 3+ in cases with > 30% positive tumor cells; **: positive if at least moderate staining intensity in > 5% of tumor cells.