The award recognises two scientists from the US and one from Germany, who developed super-resolved fluorescence microscopy.
They are Eric Betzig of the Howard Hughes Medical Institute; Stefan Hell of the Max Planck Institute for Biophysical Chemistry and the German Cancer Research Center; and William Moerner, from Stanford University.
Their ground-breaking work allowed limitations of the light microscope to be overcome, giving scientists the ability to visualise the pathways of individual molecules inside cells.
The trio brought optical microscopy into the nanodimension, according to a statement from the Royal Swedish Academy of Sciences.
In 2000 Professor Hell developed a process known as stimulated emission depletion microscopy. This uses two lasers to scan a sample and gives higher resolution images than was previously possible.
Meanwhile, working separately, Dr Betzig and Professor Moerner laid the foundations for a second method, known as single-molecule microscopy, which creates a dense super-image of nanoscale resolution. This method was used for the first time in 2006.
Nanoscopy is now used globally to produce new knowledge that is of great benefit to mankind.
None of the winners has appeared on a list of predicted Nobel winners compiled each year by Thomson Reuters citation analyst David Pendlebury.