Multiple beam interference confocal microscopy: Tool for morphological investigation of a living spermatozoon

The interference pattern obtained from a multiple internal reflection of a spermatozoon, sandwiched between the glass plate and the cover plate, was f ocused on the objective of a scanning confocal microscope. According to opt ical path differences, morphological details were revealed. The combined fe atures, namely improved resolution in z axis, originating from the interfer ence pattern and the optical sectioning of the confocal scanning system, en hanced the resolution and contrast in an impressive manner. These features permitted unprecedented images of the spermatozoon to be obtained at 0.1 mu m optical sectioning and reconstruction of three-dimensional (3-D) images. With the help of optical sectioning, it was possible to estimate the thick ness and the morphological details of the spermatozoon.