A microscope apparatus (2) for imaging a sample (5) positioned on a substrate (3) of a sample holder (1), the microscope comprising a sample stage (16) on which the sample holder is mounted, a light collecting device (38), an imaging sensor (12), and an autofocus system (8) that serves to focus an image of the sample on the imaging sensor. The autofocus system (8) is positioned below the sample stage and comprises a light beam emitter (22) emitting an autofocus light beam (13), and a beam steering system (24) configured to direct and reflect the autofocus light beam (13) off a bottom side of the substrate (3) of the sample holder positioned on the sample stage (16) and onto a light sensor (40) comprised in the autofocus system or onto the imaging sensor (12), the beam steering system configured to adjust a translational position and an angle of the autofocus light beam output by the beam steering system.