Journal: bioRxiv

Article Title: Revealing the Ultrastructure of Live Candida albicans using Stimulated Emission Depletion Microscopy

doi: 10.1101/2024.11.25.625149

Figure Lengend Snippet: (A) A comparison of the emission profile acquired using confocal laser scanning microscopy (CLSM) and STED microscopy, along with a Perrin-Jablonski diagram describing the emission depletion during the fluorescence process. A depletion beam is aligned with the excitation laser during STED imaging to deplete the emission of fluorophores in the depletion region, resulting in a smaller overall emission profile and enhanced spatial resolution. Disruption of the internal conversion process causes relaxing electrons to immediately return from the excited state (S1) to the ground state (S0), releasing a red-shifted photon which is blocked by the microscope fluorescence emission filters, while fluorescence from S1 to S0 is collected from the confined excitation volume and used to form an image. (B) A methods workflow showing the growth, staining and preparation methods for live-cell STED imaging of C. albicans .

Article Snippet: Imaging was performed using an inverted microscope equipped with confocal laser scanning microscopy (CLSM) and STED capability (STEDYCON, Abberior Instruments).

Techniques: Comparison, Confocal Laser Scanning Microscopy, Microscopy, Fluorescence, Imaging, Disruption, Staining

Journal: bioRxiv

Article Title: Revealing the Ultrastructure of Live Candida albicans using Stimulated Emission Depletion Microscopy

doi: 10.1101/2024.11.25.625149

Figure Lengend Snippet: (A) C. albicans labelled with Nile Red, imaged with a CLSM. A digitally magnified ROI is shown with a green box, confirming intracellular staining. (B) C. albicans labelled with actin LIVE 460L, imaged with a CLSM. (C) C. albicans labelled with DNA LIVE 590, imaged with a CLSM. (D), (E), and (F) show the STED counterparts of (A), (B), and (C). A digitally magnified ROI is shown in (D), showing the level of detail improvement possible with STED microscopy compared to CLSM.

Article Snippet: Imaging was performed using an inverted microscope equipped with confocal laser scanning microscopy (CLSM) and STED capability (STEDYCON, Abberior Instruments).

Techniques: Staining, Microscopy

Journal: bioRxiv

Article Title: Revealing the Ultrastructure of Live Candida albicans using Stimulated Emission Depletion Microscopy

doi: 10.1101/2024.11.25.625149

Figure Lengend Snippet: CLSM and STED microscopy of live C. albicans stained with Nile Red. (A) Representative confocal microscopy image. (B) Representative STED microscopy image of the same field of view as shown in (A). Cyan and yellow boxes highlight regions of interest (ROIs). Digitally magnified ROIs of CLSM data are shown in (C) and (E), while digitally magnified ROIs of STED data are shown in (D) and (F). Cyan arrows in (D) show mitochondria with the cristae visible, yellow arrow in (F) shows a sub-micron diameter vacuole interacting with mitochondria, and the magenta arrow in (F) highlights an example of a lipid-rich vesicle. Features indicated by arrows are not clearly visible in the corresponding CLSM micrographs (C) and (E). Scale bars for (A) and (B) = 10 µm, scale bars for (C) to (E) = 1 µm.

Article Snippet: Imaging was performed using an inverted microscope equipped with confocal laser scanning microscopy (CLSM) and STED capability (STEDYCON, Abberior Instruments).

Techniques: Microscopy, Staining, Confocal Microscopy

Journal: bioRxiv

Article Title: Revealing the Ultrastructure of Live Candida albicans using Stimulated Emission Depletion Microscopy

doi: 10.1101/2024.11.25.625149

Figure Lengend Snippet: (A) CLSM image of C. albicans labelled with Nile Red. A green line shows a ROI. (B) Line intensity profile of the ROI shown in (A). (C) Image decorrelation analysis of (A) gives a resolution of 417 nm for this CLSM micrograph. (D) STED microscopy image of C. albicans labelled with Nile Red, showing the same ROI as (A) with a magenta line. (E) Line intensity profile of the ROI shown in (D) for the STED data. (F) Image decorrelation analysis of (D) gives a resolution of 136 nm for these STED data. Scale bars for (A) and (D) = 1 µm.

Article Snippet: Imaging was performed using an inverted microscope equipped with confocal laser scanning microscopy (CLSM) and STED capability (STEDYCON, Abberior Instruments).

Techniques: Microscopy