Evaluation of DNA Fragmentation in Human Sperm by Interferometric Phase Microscopy

DNA fragmentation in sperm is considered to be one of the main reasons of the low successes rate of intracytoplasmic sperm injection (ICSI) in couples with fertility problems. Acridine orange (AO) is a dye that emits green fluorescence when binding to double-strand DNA, and red fluorescence when binding to single-strand nucleic acid. Interferometric phase microscopy (IPM) is a microscopy method that evaluates the height of different compartments in the cells.

In this work, we fixed human sperms from donors on a gridded microscope slide and captured the cells using IPM. The IPM images were analyzed by computer for the estimation of different cellular parameters. An embryologist also evaluated the IPM images. The sperm cells were than stained using AO and captured by fluorescent microscope. Each of the sperm cell was estimated by his color (red, orange, yellow and green). Both AO based and IPM-embryologist results were summarized, and statistical differences in morphological parameters between the different DNA fragmentation levels were evaluated.

Our results indicate that the size of an acrosome is a good predictor for the presents of intact DNA. We have also observed that the larger cells, in terms of cell projection and dry mass, are more likely to have intact DNA.

We conclude that IPM images for sperm prior to ICSI can help fertility technicians in selecting sperm cells with intact DNA, thus increase the chance for a successful pregnancy.