Described over 100 years ago, the Gouy phase anomaly refers to the
additional
π
phase shift that is accumulated as a
wave passes through focus. It is potentially useful in analyzing any
type of phase-sensitive imaging; in light microscopy, digital
holographic microscopy (DHM) provides phase information in the encoded
hologram. One limitation of DHM is the weak contrast generated by many
biological cells, especially unpigmented bacteria. We demonstrate here
that the Gouy phase anomaly may be detected directly in the phase
image using the z-derivative of the phase, allowing for precise
localization of unlabeled, micrometer-sized bacteria. The use of dyes
that increase phase contrast does not improve detectability. This
approach is less computationally intensive than other procedures such
as deconvolution and is relatively insensitive to reconstruction
parameters. The software is implemented in an open-source FIJI
plug-in.