Bright-field quantitative phase microscopy (BFQPM) for accurate phase imaging using conventional microscopy hardware

Abstract

Most quantitative phase microscopy methods require the use of custom-built or modified microscopic configurations which are not typically available to most bio/pathologists. There are, however, phase retrieval algorithms which utilize defocused bright-field images as input data and are therefore implementable in existing laboratory environments. Among these, deterministic methods such as those based on inverting the transport-of-intensity equation (TIE) or a phase contrast transfer function (PCTF) are particularly attractive due to their compatibility with Köhler illuminated systems and numerical simplicity. Recently, a new method has been proposed, called multi-filter phase imaging with partially coherent light (MFPI-PC), which alleviates the inherent noise/resolution trade-off in solving the TIE by utilizing a large number of defocused bright-field images spaced equally about the focal plane. Despite greatly improving the state-ofthe-art, the method has many shortcomings including the impracticality of high-speed acquisition, inefficient sampling, and attenuated response at high frequencies due to aperture effects. In this report, we present a new method, called bright-field quantitative phase microscopy (BFQPM), which efficiently utilizes a small number of defocused bright-field images and recovers frequencies out to the partially coherent diffraction limit. The method is based on a noiseminimized inversion of a PCTF derived for each finite defocus distance. We present simulation results which indicate nanoscale optical path length sensitivity and improved performance over MFPI-PC. We also provide experimental results imaging live bovine mesenchymal stem cells at sub-second temporal resolution. In all, BFQPM enables fast and accurate phase imaging with unprecedented spatial resolution using widely available bright-field microscopy hardware.

Quantitative phase microscopy via optimized inversion of the phase optical transfer function

“…In the course of this development, it is recognized and shown that the WOTF may be linearized without assuming a weakly scattering object using conditions analogous to the weak absorption and slowly varying phase conditions originally derived by Guigay [77]. Under these conditions, a new WOTF reconstruction method, which is based on an optimized inversion of the phase optical transfer function (POTF) portion of the WOTF and was briefly demonstrated in [78], is described, which enables high spatial and phase resolution using defocused bright-field micrographs without any hardware modification. Due to experimental simplicity, this algorithm, referred to as POTF recovery, may appeal broadly to end users and practitioners, thus promoting a more widespread adoption of QPI in the biomedical community.…”