Quantitative analysis of exocytosis visualized by a video‐enhanced light/fluorescence microscope reveals two distinct components of exocytosis in RBL‐2H3 cells

Abstract

Rat basophilic leukemia (RBL-2H3) cells, which exhibit Ca2+-dependent secretion of granules when stimulated with antigen or the Ca2+-ionophore A23187, were observed under a video-enhanced light/fluorescence microscope. Exocytotic events of individual granules were visualized in individual cells stimulated with antigen or A23187. Exocytosis of granules stimulated with A23187 showed two peaks in the time course. The earlier one was inhibited by selective inhibitors of protein kinase C (Ro31-8425, Ro31-8220, and chelerythrine) and the other was inhibited by an inhibitor of phosphatidate hydrolase, propranolol. Exocytosis by antigen stimulation, however, showed only one peak, which was inhibited by the selective inhibitors of protein kinase C, but not by propranolol. These results indicate that at least two distinct components of exocytosis exist in RBL-2H3 cells.

Nano-Kinetics of Probe-Particles in Solution Visualized by a Pin-Fiber Video Scope

“…7 Then, 0.50 mL of 1% sodium citrate (Katayama) was added to a boiled 50 mL portion of 0.01% chloroauric acid (Nacalai Tesque). The solution was kept boiling until it showed a wine-red color.…”

“…The g-particles obtained were pipetted on a glass slide (Mastunami Glass, 76 mm × 26 mm, 1.0 mm thick), which was cleaned with distilled water in advance. RBL-2H3 cells were incubated by a method described in a reference, 7 and DNP-BSA (Sigma, 20 ng/mL) was used as an antibody.…”

Silver nanoparticle-induced degranulation observed with quantitative phase microscopy

“…Due to the small size of vesicle (on the order of 1 μm), there is a lack of high resolution and quantitative information in light microscopy such as phase contrast and DIC. With the help of weakly basic fluorescent probes (acridine orange or quinacrine) to incorporate selectively into the low-pH granules, the secretory processes of exocytosis in mast cells could be visualized with fluorescent microscopy [17,18] . Recently, a new method combining atomic force and laser scanning confocal microscopy was developed for characterizing 3D structures of membrane and actin rearrangement during the degranulation of primary mast cell [19] .…”

Application of on-chip cell cultures for the detection of allergic response

“…6B) before addition of the stimuli. When DNP-BSA solution was introduced into the chip, the initial step of exocytosis event took place in less than 33 ms. As the solution reached the cell chamber, the exocytosis event started and continued for at least 3 min (Ozawa et al, 1996). Consequently, quinacrine release was monitored both as a PMT signal, and as the variation of the fluorescence intensity.…”