A light wave contains two primary sets of characteristics – intensity/amplitude variations and phase front variations. At present, all commercial image sensor chips are designed to operate much like our retinas and are only responsive to the intensity variations of the light wave. However, the phase front of the light wave carries additional information that may not be present in the intensity variations. For example, many biological specimens are effectively transparent and only modulate the phase front of light transmitted through them. Optical phase microscopes are greatly valued for their ability to render contrast based on refractive index variations in unstained biological samples, and are useful in biomedical applications where minimal sample preparation procedures are required. Such applications can include field analysis of bloodborne and waterborne pathogens [1
] where cost considerations and ease-of-use are important, and analysis of biopsy sections to determine tumor margins during surgical procedures where rapid processing is critical [2
]. The phase microscopes are also critical in scenarios where staining is undesirable or simply not an option. Such applications include examinations of oocytes and embryos during in-vitro fertilization procedures [3
], and longitudinal imaging of live cells or organisms [4
DIC microscopes [5
] and, to a lesser extent, phase contrast microscopes [6
] and Hoffman phase microscopes [7
] have been the primary phase microscopes of choice for the past five decades. However, the phase information is mixed with the intensity information for these phase microscopy techniques. This limitation introduces ambiguities in the rendered images and, additionally, prevents straightforward quantitative phase analysis. Moreover, these phase microscopes require special optical components that have to be switched in and out during operation. Additionally, DIC images of birefringent samples, such as muscle tissues and collagen matrices, can have significant artifacts as the DIC microscope uses polarization in its phase-imaging strategy [8
]. The relative high cost of such systems also prevents the broader use of such phase microscopes. In recent years, numerous novel phase microscopy techniques have been developed [9
]. However, the need for laser sources and the relatively high level of sophistication have thus far impeded the broader adoption of these techniques as a convenient and viable replacement for the DIC microscopes. Quantitative optical phase [12
] can also be calculated by collecting 2 or 3 successive images of the sample around its focal plane. However, this technique requires the physical actuation of the camera to be placed in distinct positions, and is therefore intrinsically limited in speed. Finally, these systems typically use relatively complex and bulky optical arrangements to translate the phase front variations into the intensity variations that are then detectable by commercial image sensor chips.
Based on our proof-of-concept experiment [13
], we believe that the implementation of a sensor chip that is capable of phase front sensing can provide a simpler and more sensible solution. Such a sensor chip can substitute for the conventional camera in a standard microscope and provide a more direct means for performing phase imaging. If such a chip can be fabricated at the foundry level, it can significantly lower the cost of phase microscopy systems and allow greater phase imaging access to the broader biomedical community.
In this paper, we report the implementation of such an image sensor chip, termed wavefront image sensor chip (WIS), that is capable of simultaneously measuring both the intensity and the phase front variations of an incident light field. The basic WIS design is closely related to the Hartmann sieve [14
] – the predecessor of Hartmann Shack sensors. Here we incorporate a grid of apertures directly on a sensor chip at close proximity to the sensor pixels. Unlike in a typical Hartmann sieve design, the WIS is able to achieve a high grid density by operating in a high Fresnel number regime.
In the Section 2, we will describe the implementation and characterization of the first fully integrated WIS prototype device. In the Section 3, we will demonstrate its capability for converting a standard microscope into a wavefront microscope (WM). In the Section 4, we will report the use of the WM for imaging polystyrene microspheres, unstained and stained starfish embryos, and strongly birefringent potato starch granules. In the Section 5, we will discuss the challenges and opportunities of the further development of the WIS. In the Section 6, we will conclude by briefly discussing the other applications of the WIS beyond enabling wavefront microscopy.