A flexible quantitative phase imaging microscope for label-free imaging of thick biological specimens using aperture masks

Abstract

A flexible quantitative phase imaging microscope is reported that offers new capabilities in terms of phase measurement from both thin and thick biological specimens. The method utilises Zernikes phase contrast approach for label-free imaging with a Twymann-Green based phase shifting module in the back focal plane. The interfering wave fronts are manipulated by laser cut apertures to form the scattered and non-scattered fields. The design is flexible and low-cost. It is shown that the bandwidth of the optical source can be optimised to enable larger optical path differences to be measured whilst giving essentially speckle free imaging. Phase maps of the cell membrane, nucleus and nucleolus of transparent epidermis cells of Allium cepa have been examined as proof of concept. Measurements from a range of glass beads confirm the optical path difference capability. The implementation of the phase shifting module is < 10% of the cost of that using a spatial light modulator whilst delivering equivalent phase resolution.