![]() At around the same time Hans Lippershey was also working on the development microscopes, as well as telescopes, and to this day the question of who was truly first remains a matter of much contention. Their design featured variable magnification and ultimately allowed objects to be magnified up to nine times. Although nobody can individually be credited for creating the first compound microscope, one of the earliest functional examples was conceived by Hans Jansen and his son, Zacharias, in the late 16th, early 17th century. The ability to see beyond the restrictions imposed by the human eye has forever changed the way we look at nature and life. Microscopy has revolutionized biological research. This review strives to provide the aspiring life science researcher with an introduction to optical microscopy, starting from the fundamental concepts governing compound and fluorescent confocal microscopy to the current state-of-the-art of super-resolution microscopy techniques and their applications. By overcoming the diffraction barrier, light microscopy could once again be established as an indispensable tool in an age where the importance of understanding life at the molecular level cannot be overstated. It should therefore not come as a surprise that the 2014 Nobel Prize in Chemistry was awarded to the scientists who, each in their own way, contributed to transforming super-resolution microscopy from a technological tour de force to a staple of the biologist's toolkit. Even though some of the fundamental concepts enabling super-resolution light microscopy have been known for quite some time, practically feasible implementations have long remained elusive. Fortunately, continuous advancements in optics, electronics and mathematics have since provided the means to once again make physics work to our advantage. The diffraction limit has restricted our view to length scales well above 250 nm and in doing so, severely compromised our ability to gain true insights into many biological systems. Check back frequently this is a rapidly developing field with regular updates to both products and publications.Ever since the inception of light microscopy, the laws of physics have seemingly thwarted every attempt to visualize the processes of life at its most fundamental, sub-cellular, level. Individual SRM techniques are highlighted below. Select the technique you're interested in, and the linked pages will help you identify the dyes that work best for that application. The Molecular ProbesĀ® product range offers multiple dye combinations that can be selected for multiplexing in any of the SRM techniques. ![]() We are a leader in fluorescence technology, and Molecular ProbesĀ® products are widely published in SRM applications. To achieve optimum performance in the various SRM techniques and more traditional two-photon microscopy, different fluorophore requirements must be met for each (details below). SRM offers techniques to approach the resolution historically reserved for electron microscopy with all the benefits of targeting and multiplexing for biological context. This means that if two or more fluorophores are within a few hundred nanometers of one another, their images become blurred together, limiting resolution. In conventional microscopy, due to interactions between visible light and the surrounding medium, a single fluorophore-which is less than a few nanometers in diameter-can only be resolved as a point spread function that is roughly half the light wavelength for lateral resolution and roughly twice that in depth. The fine mapping of cellular structures such as neural synapses, golgi apparatus, and nuclear membranes reveals more biological information when components are resolved beyond the diffraction limit and features can be elucidated at the sub-organelle level. Super-resolution microscopy (SRM) describes any optical technique used to resolve structures beyond the diffraction-limited resolution of conventional light microscopy. ![]()
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