Figure 1

(a) Schematic diagram of our optical system. A 60 × objective collects transmitted light (green) and delivers the trap laser (red). Lenses L1 and L2 are used as a beam expander to over-fill the back aperture of the 60X objective. L2 is mounted on a translation stage to allow the position of the focus and hence the trapped particle to be adjusted in z. L3 and L4 are used to create a 4f system with the camera at 4f and grating at 3f from the image plane of the microscope. (b) Schematic representation of the etch patterns of the gratings used to split images to perform multiplane microscopy. Two orthogonal gratings with etched surfaces facing each other and placed as close together as possible are necessary to view nine different focal planes.