Nomarski Imaging

As per request, I'm going to cover an application of physics today that is really on the proverbial cutting edge. Differential Interference Contrast (DIC) microscopy or Nomarski imaging is an exciting optical method that allows us to "see" microscopic, translucent biological material. As is becoming a theme, I'll need to explain a few concepts in optics before continuing to the meat of DIC imaging.

Prerequisite Light Discussion:

It's no secret that light is a rather complicated beast. First of all, a photon (basic unit) of light is simply a packet of electromagnetic radiation. Sometimes it acts like a wave (it refracts, diffracts, and reflects) and sometimes it acts like a particle (we can shoot photons one at a time, which is no more wave-like than a single water molecule by itself on the beach). To be clear, light is neither a wave nor a particle, but it acts like one or the other depending on the conditions under which we observe it. The wave part of the wave-like side of light is the behavior of the electromagnetic field of which it is composed. The electric field grows stronger and weaker with regular oscillations as does the magnetic field (oriented perpendicularly to the electric field). It is from these oscillations that we determine frequency, wavelength and other wave-like characteristics.

Polarization is the term we use to describe how all of the photons' electric and magnetic fields from a specific source are aligned. If the field oscillations in each photon have random orientations, the light is unpolarized. If all of the electric fields of each individual photon are oriented up and down, we call this vertical polarization. We can also achieve circular polarization by causing the electric fields of each photon to rotate either clockwise or counterclockwise such that at any instant, each of the photons are oriented in the same direction. This is more applicable than you might think. We use polarized filters in sunglasses to cut out reflective glare, in films to produce three dimensional effects (if you wear those silly glasses) and in astronomy (of course).


Phase is another important concept in light that we'll need to consider here. As shown in the image, two waves can be identical in amplitude, wavelength, and frequency, but can still be out of phase. This means that their moments of maximum field strength happen at different times. Phase is the reason that photographs don't look the same as real life. A picture can record the differences in intensity of light hitting the screen, but (except for in holography) film cannot record the phase difference in light adequately enough to reproduce it for the observer. The image comes out flat-looking.

DIC Imaging:

Differential interference contrast imaging uses a combination of applications in polarization and phase to image translucent images. 45-degree polarized light is split into two beams, one of 90-degree polarized light and the other of 0-degree polarized light. Though polarization is divided in this split, phase is kept constant. That means that two photons -- one 90- and the other 0-degree polarized -- that passed through the beam splitter at the same time will keep the same relative phases that they had when the beam was together. Each beam is indepentantly but simultaneously passed through the the material using a converging lens. The material is not necessarily homogenous throughout. It will have regions of high density and perhaps regions of differing composition. Since light travels a little bit slower in dense media (with a higher index of refraction), the photons passing through denser parts of the material will take a longer time to get through it. Thus, the phase of each beam becomes variable over the beam, not constant as it was at the beginning.

The beams (each now identically phase-shifted and perpendicularly polarized) are brought back together and projected onto a film. Here, you'll notice that both phase and polarization are recorded in each beam. As the beams combine, not only will they interfere (due to phase differences), causing the texture of the material to become visible through a series of brighter and darker contours, but the three-dimensionality of the material (its thickness, for example) will come out because of the polarization. To see in three dimensions, we must have two slightly different views of the same thing (such as through your left and right eyes). Polarization provides just that sort of perspective, rendering the image in three dimensions. This splitting and recombination of beams to measure objects is known as interferometry and is prevalent in optical and astronomical research, having many applications.

Thus, even though we cannot actually see the material that we are analyzing, its variable density lends itself to visible analysis by exploitation of the propensity of light to slow down in denser media. The images we get yield the kind of extreme detail required to learn about microscopic, organic materials.