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We will talk microscopy today! I need to run to the lab for a few hours, so stay tuned for a thread later! In the meantime, who has heard of super-resolution fluorescence microscopy? Or, in Greek, μικροσκοπία φθορισμού υψηλής διακριτικής ικανότητας;
Fluorescence involves the absorption of a photon by a molecule, followed by the emission of another photon. Light excites the molecule; electrons within the molecule transition from the ground state to the excited state. Upon return to the ground state the electrons emit light.
Fluorescence microscopes use this property to see molecules of interest. Several ways exist to visualise molecules by tagging them with a fluorescent label; You can also do multiple labelling and see more than one molecule at the same time!
But resolution has been the major restriction in fluorescence microscopy. Due to the diffraction limit, the discerning of structural details below approximately 200nm is impossible.
Super-resolution (SR) describes the techniques that can surpass the diffraction limit, making it possible to visualise molecules at the nm scale! This ground-breaking work was awarded the Nobel Prize in Chemistry in 2014! There are many different techniques that can achieve SR.
The SR technique I have mainly worked with is DNA-PAINT. DNA-PAINT is part of a group of techniques that utilise what is called single molecule localisation microscopy (SMLM) to achieve nm resolution.
In SMLM fluorescent emitters within a diffraction-limited area get sequentially activated and localised with sub-diffraction precision. I found this YouTube video that demonstrates that!
In DNA-PAINT, DNA strands (imagers) exhibit short-term binding to complementary docking strands, linked to a specific biological target. Imagers carry a fluorophore which provides a fluorescence signal once immobilised, leading to detection and localisation of the target.
Any questions please DM me at @annamel23 :-) Tomorrow we'll talk about the samples I am using with DNA-PAINT and what a typical week (if it all goes well first time!
) in the lab looks for me 
) in the lab looks for me
