Abstract
Correlating data from different microscopy techniques holds the potential to discover new facets of signaling events in cellular biology. Here we report for the first time a hardware set-up capable of achieving simultaneous co-localized imaging of spatially correlated far-field super-resolution fluorescence microscopy and atomic force microscopy, a feat only obtained until now by fluorescence microscopy set-ups with spatial resolution restricted by the Abbe diffraction limit. We detail system integration and demonstrate system performance using sub-resolution fluorescent beads and applied to a test sample consisting of human bone osteosarcoma epithelial cells, with plasma membrane transporter 1 (MCT1) tagged with an enhanced green fluorescent protein (EGFP) at the N-terminal.
Highlights
Correlating data from different microscopy techniques holds the potential to discover new facets of signaling events in cellular biology
Once it was verified that it is possible to work below the diffraction limit, we proceeded with correlation of the optical and Atomic Force Microscope (AFM) data
A super-resolution SIM system has been successfully integrated with a tip-scanning Quantitative Imaging (QI) nanomechanical mapping AFM for simultaneous Super-Resolved Structured Illumination Microscopy (SR-SIM)/AFM operation
Summary
Correlating data from different microscopy techniques holds the potential to discover new facets of signaling events in cellular biology. The atomic force microscope has been successfully integrated with different optical microscopy techniques[9,10,11], allowing to overcome some of its individual limitations when it comes to, for instance, sample penetration and biological specificity In this regard, AFM has been combined with Confocal Laser Scanning Microscopy (CLSM)[12,13,14], Aperture Correlation Microscopy[8], Total Internal Reflection Fluorescence Microscopy (TIRFM)[15,16,17,18,19,20,21], and Fluorescence Lifetime Imaging (FLIM)[22,23]. Recent research has proposed a new dye to circumvent this problem that allows correlative AFM and STORM imaging without the need to change the buffer[30]
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