Journal: EMBO Reports

Article Title: Detecting material state changes in the nucleolus by label-free digital holographic microscopy

doi: 10.1038/s44319-024-00134-5

Figure Lengend Snippet: Description of the beam path and microscope configuration used. The diagram illustrates the detailed beam path in our QMOD setup. A purposely built versatile “plug-in” DHM adapter (QMOD, developed together with Ovizio s.a.) was connected between the lateral port of a Zeiss inverted microscope and a Retiga R3 camera (Qimaging) used for imaging the DHM phase and all fluorescence channels. The camera was driven from the MetaXpress (Molecular Devices) environment. Images were processed with OsOne (Ovizio). Digital holograms were recorded with an incoherent light source ( HAL lamp) on an inverted microscope adjusted for proper Köhler illumination and coupled to a QMOD interferometer and CCD camera. A bandpass filter ( BP , 550 nm) was used to increase the coherence of the light and obtain the partial coherence required for holography. The image-forming light rays passing through the specimen ( S ) were captured with the microscope objective ( O ) and directed from microscope lens L1 to the QMOD interferometer. In the QMOD, a diffraction grating G induced splitting of the incident light beam into a diffracted beam ( 1 , reference) and a non-diffracted light beam ( 2 , object beam). A second lens ( L2 ) placed at the focal distance from the grating G reshaped both the diffracted and non-diffracted beams into beams parallel to the optical axis. A wedge ( W ) inserted in the optical path of the object beam induced a slight shift of the images produced by the diffracted and non-diffracted light beams. C is a compensating optical module placed in the optical path of the non-diffracted light beam to compensate for the light shift introduced by W in the diffracted beam. The diffracted beam is then recombined with the object beam and focalized by means of objective lens L3 on the recording plane of a CCD camera, where the hologram is recorded. LED, illumination; FF, fluorescent filter cube. Fluorescence imaging: excitation illumination is emitted by a light-emitting diode ( LED ) and is directed to the sample ( S ) through a fluorescence filter cube ( FF ). Fluorescence emission by the specimen is collected by the objective, passes through the filter cube and L1 , enters the QMOD, and finally reaches the CCD camera.

Article Snippet: Single-band bandpass optical filter , Semrock , FF01-550/49-25.

Techniques: Microscopy, Inverted Microscopy, Imaging, Fluorescence, Produced

Journal: EMBO Reports

Article Title: Detecting material state changes in the nucleolus by label-free digital holographic microscopy

doi: 10.1038/s44319-024-00134-5

Figure Lengend Snippet: Reagents and tools table

Article Snippet: Single-band bandpass optical filter , Semrock , FF01-550/49-25.

Techniques: Recombinant, Sequencing, Membrane, Plasmid Preparation, Software