Auto-focus is an important technique for auto reconstruction of digital holograms, and works by focusing criterion function which influences the effect of the autofocus greatly. Aiming at Marine Microorganism holograms, this paper proposes a new focused criterion function established according to characteristics of detail discrete degree in reconstruction image changing with FD. Simulation results illustrate that, the proposed method, compared with several existing methods, has better single peak, higher sensibility and higher signal to noise ratio (SNR). The new method can realize autofocus of reconstruction distance to implement auto reconstruction of holograms.