Normalizing projection images: a study of image normalizing procedures for single particle three-dimensional electron microscopy

In the process of three-dimensional reconstruction of single particle biological macromolecules several hundreds, or thousands, of projection images are taken from tens or hundreds of independently digitized micrographs. These different micrographs show differences in the background grey level and particle contrast and, therefore, have to be normalized by scaling their pixel values before entering the reconstruction process. In this work several normalization procedures are studied using a statistical comparison framework. We finally show that the use of the different normalization methods affects the reconstruction quality, providing guidance on the choice of normalization procedures.