Abstract

Quantitative imaging refers to imaging where local image intensities can be directly related to the abundance of markers and, ideally, targets in biological samples. This has become especially relevant with the advent of molecular resolution imaging in the form of fluorescence super-resolution microscopy. In this talk I will report on our efforts towards achieving truly quantitative and molecular resolution microscopy of ion channels in single cells. Recent advances have been enabled by progress in several areas, including improved camera technology, image analysis and marker approaches. An important role for DNA nanotechnology has arisen from the ability to create nanostructures that are ideally suited to assessing imaging performance and providing calibration standards. I will illustrate how this can be used in routine super-resolution microscopy for quality assurance and algorithm testing. Perhaps even more importantly, the use of synthetic DNA can be used to control the transient binding of dye molecules to markers, known under acronyms such as DNA -PAINT. This approach has provided enormous flexibility in designing labelling schemes compatible with high resolution and accurate quantification. These advantages will be illustrated with the true molecular scale visualization of variable clustering properties of ryanodine receptors, a large calcium channel, in cardiac myocytes.