Evaluation of the fluorescein diacetate (FDA) vital dye viability test with hamster and bovine embryos

Abstract

A study was undertaken to determine: (1) the potential toxicity of a fluorogenic vital dye, fluorescein diacetate (FDA), on hamster and bovine pre-implantation embryos; and (2) whether a correlation exists between the fluorescence of an embryo and its ability to continue development. For the toxicity trial, hamster eight-cell embryos were randomly assigned to one of three groups (control, FDA+UV light or UV light only), and early bovine blastocysts to either a control or FDA+UV light group. Embryos were cultured for 24 h and scored for development to the blastocyst stage. Embryos of both species developed equally well in vitro regardless of whether or not they had been exposed to FDA and UV light. Treated and untreated control embryos from both species were transferred to synchronized recipients. Similar numbers of pregnancies resulted after transfer of treated and untreated embryos from both species. In the second experiment, the proportions of fluorescing embryos were compared using two groups of hamster eight-cell embryos: (1) freshly collected embryos; and (2) cultured embryos that failed to develop. Significantly more of the fresh eight-cell embryos fluoresced than did the cultured, undeveloped embryos. No false negative results were obtained (embryos that developed but failed to fluoresce). However, approximately half of the non-developing, cultured embryos showed varying degrees of fluorescence (false positive). Embryos showing “false positive” fluorescence may be viable, but incapable of further development due to deficiencies of the culture medium. The procedures used in the FDA viability assay were not detrimental to development of late cleavage stage mammalian embryos and thus seem suitable for rapid screening of manipulated embryos for potential damage. However, further work is needed to establish the significance of the false positive results encountered in this study.