Immunofluorescence Assay: A Method to Identify Tumor Cells Captured on a Medical Wire

- Take a functionalized wire having a small gold portion coated with a hydrogel coupled with antibodies of interest. Dip the gold part of the wire in a vial containing tumor cells. Close it with a wire stopper, and incubate it in a tube rotator. The circulating tumor cells or CTCs displaying surface antigens bind to the antibodies on the wire. Rinse the wire by dipping in vials containing phosphate buffered saline or PBS and air dry. Now dip the wire in an acetone solution to fix the cells by dehydrating them.

Next, gently insert the wire through a P200 tip with the functionalized part within it. Fill the tape with an antibody conjugated with a fluorochrome to submerge the wire. These labeled antibodies will bind to their respective antigens present on the cells. Seal the opening with a laboratory film and incubate overnight in dark at 4 degrees Celsius. Next, wash the wire with PBS to remove any unbound antibodies. Place the wire into a wire holder and view under a fluorescent microscope. The CTCs bound by antibodies fluoresce on the wire. In the following protocol, we will capture CTCs on a medical wire and identify them by immunofluorescent staining.

- To begin the experiment, r