Automatic numerical focus plane estimation in digital holographic microscopy using calibration beads

We present a new method to achieve autofocus in digital holographic microscopy. The method is based on inserting calibrated objects into a sample placed on a slide. Reconstructing a hologram using the inverse problems approach makes it possible to precisely locate and measure the inserted objects and thereby derive the slide plane location. Numerical focusing can then be performed in a plane at any chosen distance from the slide plane of the sample in a reproducible manner and independently of the diversity of the objects in the sample.