Highly stable digital holographic microscope using Sagnac interferometer

Abstract

Interferometric microscopy has grown into a very potent tool for quantitative phase imaging of biological samples. Among the interfermetric methods, microscopy by digital holography is one of the most effective techniques, especially for studying dynamics of cells. Imaging of cell fluctuations requires digital holographic setups with high temporal stability. Common path setups in which the object and the reference beams encounter the same set of optical elements provide better temporal stability compared to two-beam setups. Here, we present a compact, easy-to-implement, common path digital holographic microscope based on Sagnac interferometer geometry. The microscope is implemented using a diode laser module employing a CCD array or a webcam sensor to record holograms. The system was tested for three-dimensional imaging capability, numerical focusing ability, and temporal stability. Sub-nanometer temporal stability without external vibration isolation components was obtained in both cases. The higher temporal stability makes the microscope compatible to image cell fluctuations, which is demonstrated by imaging the oscillation of the cell membrane of human red blood cells.

Quantitative phase imaging based on Fresnel diffraction from a phase plate

“…In the dynamic measurements, different environmental effects between the beams, namely, the individual mechanical vibration of optical components, inevitably trigger some remnant time-dependent noises which reduce the quality of * m-dashtdar@sbu.ac.ir retrieved images. Recently, many efforts have been devoted toward developing new settings with the higher phase sensitivity such as self-reference or common-path interferometers (CPIs), through which the phase stability of the system is significantly enhanced as the two overlapping beams travel along nearly equivalent paths [12][13][14][15][16][17][18][19][20][21][22][23][24][25][26]. Despite having the several advantages including simplicity and phase stability, many of the CPIs suffer from functional challenges which are imposed by loss of flux, inflexible off-axis angle, small field of view (FOV) limitation and precise optical alignment.…”

Wide field of view common-path lateral-shearing digital holographic interference microscope

“…[19][20][21][22][23][24][25][26][27][28][29][30][31][32][33][34][35][36][37][38] Common-path techniques duplicate the object wavefront and generate the reference wavefront from one of these duplicated wavefronts. This can be done by either spatially filtering one of the duplicated wavefronts and using it as the reference wavefront [19][20][21][22][23][24][25][26][27][28][29] or using a portion of the duplicated wavefront (which does not contain object information) as the reference wavefront. [30][31][32][33][34][35][36][37][38] A second technique termed a self-referencing DHIM technique requires fewer optical elements and hence leads to very compact off-axis geometries.…”

“…24 One of the advantages of the common-path DHIM is that it can provide subnanometer temporal stability which is required to study cell membrane fluctuations. [19][20][21][22][23][24][25]28,[35][36][37] Consequently, the measurement of temporal stability of the developed common-path system is necessary before its use in dynamic imaging of cells. For measurement of the temporal phase stability, a time series of holograms with a blank microscope slide (object holograms) in the sample holder is recorded.…”

“…The mean of the standard deviations of the time variation of the phase (obtained from hologram recorded at each time instance) computed at several spatial points provides the temporal phase stability. [23][24][25]28,[35][36][37] This is repeated by recording several time series, and the average of these phase stabilities is used as the value of the temporal phase stability of the system. For the CCD array, holograms were recorded at the rate of 15 Hz for a total period of 30 s, and, in the case of the webcam sensor, holograms were recorded at 25 Hz for 30 s to compute the temporal stability.…”

Nanoscale surface characterization using laser interference microscopy

“…This class brings together methods of white light interferometry [1,2], phase shift interferometry [3][4][5], digital holographic microscopy [6,7], laser interference microscopy [8,9], coherent phase microscopy [10], coherent correlation interferometry [11,12], etc.…”