Focal Spots of Size <mml:math xmlns:mml="http://www.w3.org/1998/Math/MathML" display="inline"> <mml:mi>λ</mml:mi> <mml:mi>/</mml:mi> <mml:mn>23</mml:mn> </mml:math> Open Up Far-Field Florescence Microscopy at 33 nm Axial Resolution

Abstract

We report spots of excited molecules of 33 nm width created with focused light of lambda = 760 nm wavelength and conventional optics along the optic axis. This is accomplished by exciting the molecules with a femtosecond pulse and subsequent depletion of their excited state with red-shifted, picosecond-pulsed, counterpropagating, coherent light fields. The lambda/23 ratio constitutes what is to our knowledge the sharpest spatial definition attained with freely propagating electromagnetic radiation. The sub-diffraction spots enable for the first time far-field fluorescence microscopy with resolution at the tens of nanometer scale, as demonstrated in images of membranes of bacillus megaterium.