Figure 1

Detection of CK19 protein. (a) CK19-EPISPOT assay procedure. Day 1: The membranes of the ELISPOT plates are coated with an anti-human CK19 monoclonal antibody (mAb, Ks19.1). Days 2 to 3: Next, the cells are seeded in each well and cultured for 48 hours. During this incubation period, the released specific proteins are directly immunocaptured by the immobilized mAb on the bottom of the well. Plates are then washed, and cells are removed. The presence of the released CK19 protein is revealed by the addition of an anti-human CK19 mAb (Ks19.2) conjugated with a fluorochrome. Day 4: Fluorescent immunospots are counted with an automated reader. One immunospot corresponds to the fingerprint left only by one viable releasing epithelial tumor cell. (b) Schematic representation of the assumed binding sites of anti-cytokeratin antibodies and cleavage site for caspase 3 on the human CK19 protein [3, 40].