Live Cell Imaging of Human Mast Cells on a Pico-well LiveCell Array at Single Cell Resolution (B207)

Abstract

Mast cells are important effecter cells in allergic and inflammatory responses. When the mast cells are activated they undergo regulated secretion of intracellular granules by exocytosis. This includes inflammatory cytokines such as IL-6, TNF-α and other chemokines. In this study we examined mast cell activation using a microscope slide based pico-well array at a single cell resolution. The human umbilical cord blood derived mast cells were cultured in the mast cell medium. The cells were counted and 2uL of cell suspension were added to the pico-wells of the array (Livecell array, NUNC). The cells in the array were covered with a sliding cover slip allowing each cell to settle into an individual well. The cover slip also allowed for repeated treatment and washing of reagents via capillary flow. The cells were treated with substance P (SP) to induce activation directly on the array. Live cell imaging showed cells were distributed evenly on the array as one cell per well. After activation, the cells secreted granules which are clearly visible in the pico-wells surrounding the cells. Additionally, LAD2 cells were activated with compound 48/80 (0.1ug/ml). Use of anti-CD63 PE permitted single cell investigation of degranulation. The ability to culture, treat and image non-adherent mast cells in the array allowed us to study single cell responses in a mast cell population, and permits fast screening of potential inhibitors.