Differential interference contrast (DIC) microscopy, also known as Nomarski interference contrast microscopy, is an optical imaging technique used in microscopy to enhance the contrast of transparent and colorless specimens. DIC microscopy provides three-dimensional, relief-like images by exploiting the interference of polarized light.
Key features and principles of differential interference contrast microscopy include:
Polarized light: DIC microscopy uses polarized light to enhance image contrast. Polarizers and beam splitters are employed to create two mutually perpendicular light beams that interact with the specimen.
Nomarski prisms: A pair of Nomarski prisms, also known as Wollaston prisms, is a key component of DIC microscopy. These prisms split the incident light into two beams with a slight lateral displacement.
Shearing interference: The two split beams pass through the specimen and are then recombined by a second pair of Nomarski prisms. The slight lateral displacement results in a shearing interference pattern, where changes in specimen thickness and refractive index produce optical disparities.
Optical path difference (OPD): The shearing interference pattern generates an optical path difference that is proportional to the gradient of the specimen's refractive index. This gradient information provides contrast for structures that are otherwise challenging to visualize, such as cell boundaries and organelles.
Relief-like images: The interference patterns create a relief-like appearance in the images, allowing for the visualization of subtle variations in specimen thickness and refractive index.
Three-dimensional appearance: DIC microscopy provides a three-dimensional appearance to specimens, enhancing the perception of depth and surface features.
Applications: DIC microscopy is widely used in biological and biomedical research, as well as materials science, to study live cells, tissues, and transparent specimens. It is particularly valuable for imaging unstained samples, providing detailed views of cellular structures.
Mitigation of halos: DIC microscopy is known for producing images with reduced halo and diffraction artifacts compared to other contrast-enhancing techniques. This contributes to clearer and more accurate representations of specimens.
No need for staining: One of the advantages of DIC microscopy is that it allows for the observation of live, unstained specimens. This is beneficial for preserving the natural state of biological samples without introducing artifacts associated with staining procedures.