Digital holographic microscopy (DHM) and transport of intensity (TIE) phase imaging of live cells

摘要

Quantitative phase imaging (QPI) provides a label free method for imaging live cells and allows quantitative estimates ofcell volume. Because the phase of light is not directly measurable at an imaging sensor, QPI techniques involve bothhardware and software steps to reconstruct the phase. Digital holographic microscopy (DHM) is a QPI technique thatutilizes an interferometer to combine a reference beam with a beam that passes through a specimen. This produces aninterference pattern on the image sensor, and the specimen’s phase can be reconstructed using diffraction algorithms.One limitation of DHM is that the images are subject to coherent diffraction artifacts. Transport of intensity (TIE)method, on the other hand, uses the fact that defocused images of a specimen depend on the specimen’s phase todetermine the phase from two or more defocused images. Its benefit over DHM is that it is compatible with conventionalbright field imaging using sources of relatively low coherence. Although QPI methods can be compared on a variety ofstatic phase targets, these largely consist of phase steps rather than the phase gradients present across cells. In order tocompare the QPI methods described above on live cells, rapid switching between QPI modalities is required. We presentresults comparing DHM and TIE on a custom-built microscope system that allows both techniques to be used on thesame cells in rapid succession, which allows the comparison of the accuracy of both measurements.