Methods have been developed to map the distribution of elements in cellular organelles and large macromolecular assemblies using electron-spectroscopic imaging in a scanning transmission electron microscope (STEM) equipped with a cold field-emission gun and a magnetic sector electron spectrometer. To optimize the elemental detection limits of the spectrometer, we have now replaced the photodiode detector with a cooled CCD detector. Advantages of this new system are (i) the low noise level that provides higher analytical sensitivity, (ii) the short read-out time of around 10 ms, which provides faster acquisition, (iii) the two-dimensional array of square pixels, which allows exact correction for channel gain variations, and (iv) the improved point-spread function of the optical coupling of the detector array to the scintillator. It was found that the root mean square read-out noise of the detector was only 3 primary electrons per spectral channel, which corresponds to a detective quantum efficiency of about 0.5 for 10 primary electrons. The system has been applied to map calcium in freeze-dried cryosections of sympathetic neurons. It was demonstrated that elemental maps containing 100,000 pixels could be acquired with sufficient counting statistics to detect physiological levels of calcium. Measurements also indicate that the CCD detector should be capable of mapping single atoms of phosphorus using the spectrum-imaging technique with a nanometer diameter probe.
