Optimal resolution in Fresnel incoherent correlation holographic fluorescence microscopy

Abstract

Fresnel Incoherent Correlation Holography (FINCH) enables holograms and 3D images to be created from incoherent light with just a camera and spatial light modulator (SLM). We previously described its application to microscopic incoherent fluorescence wherein one complex hologram contains all the 3D information in the microscope field, obviating the need for scanning or serial sectioning. We now report experiments which have led to the optimal optical, electro-optic, and computational conditions necessary to produce holograms which yield high quality 3D images from fluorescent microscopic specimens. An important improvement from our previous FINCH configurations capitalizes on the polarization sensitivity of the SLM so that the same SLM pixels which create the spherical wave simulating the microscope tube lens, also pass the plane waves from the infinity corrected microscope objective, so that interference between the two wave types at the camera creates a hologram. This advance dramatically improves the resolution of the FINCH system. Results from imaging a fluorescent USAF pattern and a pollen grain slide reveal resolution which approaches the Rayleigh limit by this simple method for 3D fluorescent microscopic imaging.

Incoherent digital holography with phase-only spatial light modulators

“…Finally, reducing the optical path difference (OPD) in FINCH, for processing wide-bandwidth light sources, was done by inefficiently increasing the length of the systems. These difficulties were fruitfully solved in the later versions of FINCH; 19,21,24 some of them are reviewed in Sec. 2.2.…”

Multispectral hyperbolic incoherent holography

“…Various types of techniques have proposed. One technique is based on diffraction theory which can be implemented by making use of spatial light modulator [3][4][5] or concave mirror [6] and other technique uses radial shearing [7] or rotational shearing [8] interferometric systems.…”

“…In Sect. 4, we investigate 3D imaging properties of hyperbolic holography by making use of a novel analytical solution of IRF defined over the 4D space. The primary result obtained in this section is a full description of the 4D IRF expressed mathematically in closed form.…”

Single channel in-line multimodal digital holography

“…The SLMs are placed with their active axes perpendicular to each other and at a 45°angle to the transmission axis of the two polarizers. The second polarizer is used to isolate, from the two orthogonal polarizations, two polarization components that have the same polarization direction and can, therefore, interfere [9]. As the input object is illuminated with a collimated laser beam, each SLM only phase-modulates components of the incident light The two independent beams are signified by red and blue.…”