eclipse ti inverted a1r laser scanning confocal microscope  (Nikon)

Journal: Nature Communications

Article Title: Confocal interferometric scattering microscopy reveals 3D nanoscopic structure and dynamics in live cells

doi: 10.1038/s41467-023-37497-7

Figure Lengend Snippet: a The main components of a confocal laser scanning microscope (CLSM), which is extended to perform iSCAT microscopy in both wide-field and confocal modes. OBJ objective, BS beam splitter, PH pinhole, DC dichroic mirror, EF emission filter, PMT photomultiplier tube. Inset: Wavefronts of laser illumination (dashed lines) and sample radiation (solid lines) for the three modalities. Lateral and axial point spread functions (PSF) of a 100 nm fluorescence-labeled polystyrene bead for wide-field iSCAT ( b , c ), confocal iSCAT ( d , e ), and confocal fluorescence ( f , g ) modalities. The focus was scanned over 4 μm in steps of 30 nm in c , e , and g . The background was accounted for in each z plane. Curves on the right-hand side depict the intensity profiles along the cross sections shown in each figure. Horizontal and vertical scale bars are 200 nm and 500 nm, respectively. h Scanning electron micrograph of a nanofabricated test sample consisting of two chromium pillars of diameter 45 nm, height 45 nm and center-to-center separation 130 nm. Scale bar is 200 nm. i C-iSCAT image of the sample recorded with a pinhole setting of 0.3 AU at a wavelength of 445 nm. Scale bar is 200 nm. j Cross sections along the white dotted lines from ( h , orange) and ( i , blue). The green curve also shows a cross section from a C-iSCAT image recorded with a pinhole setting of 1.2 AU. k – n The plasma membrane of a HeLa cell simultaneously imaged in W-iSCAT ( k ), C-iSCAT ( l ) and confocal fluorescence ( m ) modes. The plasma membrane was fluorescence-labeled with GFP-GPI. The W-iSCAT image is flat-fielded, whereas the C-iSCAT image is presented in its raw form. n An overlay of the images in l and m . Scale bars in k – n are 2 μm.

Article Snippet: Figure a shows the schematics of an inverted confocal laser scanning microscope (CLSM, Nikon Ti Eclipse A1R) that was modified to allow for iSCAT imaging in wide-field (W-iSCAT) and confocal (C-iSCAT) modes in addition to confocal fluorescence (CF) microscopy (see Methods).

Techniques: Laser-Scanning Microscopy, Microscopy, Fluorescence, Labeling, Clinical Proteomics, Membrane