Video‐enhanced contrast polarization (AVEC‐POL) microscopy: A new method applied to the detection of birefringence in the motile reticulopodial network of allogromia laticollaris

Abstract

A new method is described for recording rapid processes of cell motility in polarized light. The Allen video-enhanced contrast (AVEC-POL) method of polarization microscopy achieves significant improvements in resolution, contrast, and the visibility of fine detail by a combination of novel adjustments to a standard (unrectified) polarizing microscope and video camera. Using the full working aperture of a high-power planapochromatic objective lens and compensator setting of λ/9-λ/4, visible images appear lacking in contrast. However, the same images viewed with an appropriate video camera equipped with an electronic offset adjustment can be made to appear with as much contrast as desired, revealing a significantly greater amount of fine detail in the image than can be seen by high extinction visual microscopy alone. At bias retardations between one-ninth and one-quarter wave, the diffraction anomaly observed near extinction disappears. Consequently, polarizing rectifiers are not required with the AVEC-POL method, and images previously requiring photographic exposures of around 20 seconds are sufficiently bright to be registered on the video monitor in 1/60 second. Using an intensity monitor, quantitative measurements of cellular birefringence can be retrieved from live or videotaped images displaying a linear relationship between contrast and phase retardation due to birefringence. The AVEC-POL method also renders accessible to polarized light analysis a number of objects that scatter or depolarize too much light to be studied by high extinction methods. The method is demonstrated on model objects and applied to the highly motile reticulopodial network of Allogromia laticollaris. Rapid motion in close association with microtubules can now be analyzed in greater detail at a significant reduction in the cost of recording.