SLIM Overview

Real-time, nondestructive SLIM (see Fig. 1) uses optical interferometry to reveal structure and dynamics with nanometer sensitivity in live cell cultures in 4D (3D time-series).51,52,35 In PC illumination, light beams emanating from the condenser ring pass through the sample and are collected in the back focal (Fourier) plane of the microscope objective.53 This scattered beam light is geometrically separated from the reference beam in the Fourier plane, and combined through interference in the image plane.

A phase plate in the Fourier plane advances the phase of the reference beam by 0.5π radians and attenuates its amplitude. Destructive interference generates a dark image for the dense portions of the cell with respect to the gray background. The SLIM module includes a one-to-one 4f relay system with a spatial light modulator (SLM) in the Fourier plane, and relays the microscope image plane with minimal aberrations (diffraction-limited) at a 1:1 ratio to a camera placed at its exit port. A phase mask ring, projected on the SLM screen at the same size, is conjugated with the back focal plane of the microscope objective. The SLM thus modulates the reference beam like a phase plate with variable thickness.

To create a quantitative phase image, the SLM shifts the phase of the reference beam by a fixed amount (0, 0.5π, π, 1.5π) and the camera captures the intensity image. A quantitative phase image is uniquely determined by combining the four frames and solving the field interference equations at each pixel. SLIM has successfully measured cell cycle-dependent growth and kinetics,54,55 enabled study of red blood cell structure,56 characterized the 3D structure of unlabeled live cells,57,48 and facilitated development of label-free cancer markers.58,59