- For CLSM, specimens can be examined using any appropriate
confocal laser scanning microscope (Fig.2b) and fluorescence
staining intensity can be quantified using the Image J software
(NIH). - For nanoscale visualization, STED microscopy can achieve
resolutions of 5–10 times higher than that which is capable
by CLSM (approximately 20–40 nm), allowing the capture of
very fine structural details (Fig.3). For high resolution imag-
ing, the samples can be examined on a STED nanoscope system
in conjunction with a 100oil immersion objective. This
advanced imaging technique is based on CLSM and achieved
by switching fluorophores on and off in time [18]. Analysis and
quantification of STED microscopy images can be performed
using the appropriate image analysis software.
Fig. 3High resolution imaging of scaffold IQGAP1 in 3O-C 12 -HSL-treated human
epithelial Caco-2 cells. Cell monolayers were stimulated with 12μM 3O-C 12 -
HSL for 20 min. Cells were fixed and stained with mouse anti-IQGAP1 and Atto
647 N goat anti-mouse antibodies and visualized by confocal microscopy (upper
image, inred) and STED nanoscopy (bottomimage, ingreen). Bar: 1μm
Autoinducer Effects on Mammalian Cells 223