Encyclopedia of Experiments: Cancer Research
JoVE 비디오를 활용하시려면 도서관을 통한 기관 구독이 필요합니다. 전체 비디오를 보시려면 로그인하거나 무료 트라이얼을 시작하세요.
Transcript
Under pathological conditions like cancer, activated neutrophils release neutrophil extracellular traps or NETs - a mesh of extracellular DNA with associated enzymes and antimicrobial proteins - that entrap circulating cancer cells.
To visualize the adhesive interaction between NETs and tumor cells in vitro, begin by culturing low-density neutrophils, a neutrophil subtype, in a polymer-coated culture dish that promotes cell attachment. Incubate for a few hours to stimulate the neutrophils to form web-like NETs.
Add a suspension of red fluorescein-labeled cancer cells to the neutrophil culture. Incubate the cancer cells with NETs for a brief period to facilitate cancer cell-NET interaction. The NET-DNA acts as a chemotactic factor that attracts cancer cells and promotes their adhesion to NETs.
Remove the spent medium. Gently wash the culture with a suitable fresh medium to remove any unattached tumor cells from the culture well.
Now, add a membrane-impermeable green fluorescent nuclear stain to color the extracellular NETs selectively. Observe the culture under a fluorescence microscope. Neutrophil extracellular traps appear as green string-like structures that form a mesh to which red tumor cells are attached.