EMBRYOFREEZE™

Generally, in the first years of slow freezing, DMSO was used for cryoprotection. Indeed, early cleavage stage embryos can be frozen in either 1,2-propanediol (PROH) and DMSO, both returning comparable satisfactory results (Shaw, et al. 2000). Nowadays, DMSO is successfully replaced by PROH because of lower toxicity. Thus, in most cases, with very few exceptions, a combination PROH and sucrose is used for the cryopreservation of early embryos (Mandelbaum and Ménézo 2001) (Veeck, et al. 2009).

PROH can readily permeate cell membranes and establish hydrogen bonds with water molecules to prevent ice crystallization. Consequently, PROH causes a slight lowering of the freezing point of the solution. Also, during freezing, the free water solidifies into ice and remaining solution will contain progressively higher concentration of electrolytes, reaching their toxicity level. The cryoprotective agent has the ability to reduce toxic effects of high concentrations of solutes, including salts. The protection may originate from the ability of PROH to replace water molecules around compounds, such as enzymes, hereby protecting them from irreversible conformational changes, which may occur at high salt concentrations (Chang, et al. 2010). 

Cryoprotective solutions have been supplemented with macromolecules such as sucrose and human serum albumin (HSA). Several studies have shown that such molecules help to reduce physical damage and help to maintain osmotic pressure of the extracellular fluid (Shaw et al. 2000).

Besides its cryoprotective role, HSA facilitates gamete or embryo manipulation by preventing adsorption to the surface of petri dishes and pipettes through saturation of the potential binding sites.