Cells were fixed, and prepared for immunocytochemistry staining following the protocol in Jackson et al.17 (link). Antibodies and concentrations are listed in Table 2 . Images were captured at random positions using an inverted epi-fluorescence microscope (Nikon Eclipse Ti with a DS-Qi1 camera; Nikon Instruments, Tokyo, Japan) at 200X magnification. The exposure length and gain were kept constant. ImageJ software was used to process the images. The percentage of positive staining for each marker was calculated based on an average from counting ~100 cells from randomly selected positions in n = 4 wells. Counts were verified by two independent investigators.
Antibodies Used in Immunocytochemistry.
| Antibody | Epitope | Supplier | Concentration |
|---|---|---|---|
| Proliferating Cell Nuclear Antigen (PCNA) | PC10 (mouse) | Dako | 1:500 |
| Cleaved Caspase-3 | D175 (rabbit) | Cell Signaling | 1:400 |
| Tumor Protein P63 (P63) | EPR5701 (rabbit) | Abcam | 1:300 |
| Tumor Protein P63 (P63) | 4A4 (mouse) | Abcam | 1:100 |
| ABC Transporter Family G2 (ABCG2) | BXP21 (mouse) | Santa Cruz | 1:100 |
| CCAAT/enhancer-binding protein delta (C/EBPδ) | Polyclonal (rabbit) | Abcam | 1:600 |
| Cytokeratin 14 (CK14) | LL002 (mouse) | Abcam | 1:300 |
| Cytokeratin 10 (CK10) | Polyclonal (rabbit) | Abcam | 1:800 |
| Involucrin | SY5 (mouse) | Santa Cruz | 1:300 |
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