Light microscopy imaging is being transformed by the application of computational methods that permit the detection of
spatial features below the optical diffraction limit. Successful localization microscopy (STORM, dSTORM, PALM,
PhILM, etc.) relies on the precise position detection of fluorescence emitted by single molecules using highly sensitive
cameras with rapid acquisition speeds. Electron multiplying CCD (EM-CCD) cameras are the current standard detector
for these applications. Here, we challenge the notion that EM-CCD cameras are the best choice for precision localization
microscopy and demonstrate, through simulated and experimental data, that certain CMOS detector technology achieves
better localization precision of single molecule fluorophores. It is well-established that localization precision is limited
by system noise. Our findings show that the two overlooked noise sources relevant for precision localization microscopy
are the shot noise of the background light in the sample and the excess noise from electron multiplication in EM-CCD
cameras. At low light conditions (< 200 photons/fluorophore) with no optical background, EM-CCD cameras are the
preferred detector. However, in practical applications, optical background noise is significant, creating conditions where
CMOS performs better than EM-CCD. Furthermore, the excess noise of EM-CCD is equivalent to reducing the
information content of each photon detected which, in localization microscopy, reduces the precision of the localization.
Thus, new CMOS technology with 100fps, <1.3 e- read noise and high QE is the best detector choice for super resolution precision localization microscopy.