Optisol gs

Condition A was endothelium-flapped inwards and tissue preserved in organ culture medium supplemented with 6% dextran T500 (transport medium) and shipped at room temperature (RT) (figure 1A). For condition B, the endothelium was scrolled outwards in a modified Jones tube preserved in hypothermic medium (Optisol-GS, Bausch & Lomb, Bridgewater, NJ, USA) at 4°C during transportation (figure 1B).

The ethical approval for study protocol was received from the institutional review board. All procedures conformed to the tenets of the Declaration of Helsinki.
Human corneoscleral buttons with consent for research use were obtained from Miracles of Sight Eye bank, North Carolina, USA. Tissue processing and experimentation were performed by a single surgeon (MB) with experience of >100 DMEK tissue preparations. All tissue had a central EC count of >2200 cells/mm² and had been stored in Optisol GS (Bausch & Lomb, Rochester, New York, USA) for no longer than 14 days prior to experimentation. Initial cell density measurement was performed by specular microscopy in the eye bank supplying the tissue. Prior to experimentation, specimens were assessed using 0.4% trypan blue (Sigma, St. Louis, Missouri, USA) and excluded if large areas of cell loss were visible.
Samples were assigned to one of two methods of DMEK preparation (n=8 in each group). When paired tissue was available, one cornea from each pair was assigned to each intervention. Unpaired tissue was assigned alternately.

PBS, HBSS, DMEM/F12 (Thermo Fisher Scientific, Inc.), UW solution (Viaspan^®; Bristol-Myers Squibb, Co.), ET-Kyoto solution (Otsuka Pharmaceutical Factory, Inc.), and Optisol GS™ (Bausch & Lomb, Rochester, NY) were prepared as candidates for a basal preservation medium. The following additives were also prepared: dextran 40 (0.2, 2, 20 g/L; Wako), chondroitin sulfate A (0.01, 0.1, 1%; Sigma-Aldrich, St. Louis, MO, USA), N-acetylcysteine (0.1, 1, 10 mM; Wako), allopurinol (0.1, 1, 10 mM; Tokyo Chemical Industry Co., Ltd), glutathione (0.3, 3, 30 mM; Sigma-Aldrich), adenosine (0.05, 0.5, 5 mM; Sigma-Aldrich), hyaluronic acid (0.01, 0.1, 0.5%; Calbiochem), dibutyryl cAMP (0.2, 2, 20 mM; Enzo Life Sciences), trehalose (12, 120, 1200 mM; Hayashibara), tert-butylhydroquinone (tBHQ; 0.1, 1, 10 μM; Sigma-Aldrich), oltipraz (1, 10, 100 μM; Sigma-Aldrich), and ebselen (1, 10, 100 μM; Sigma). Each of these additives was added to HBSS individually.

This study was approved by Singhealth centralized institutional review board (Ref: 2016/2839). All research-grade human cadaver corneal tissues were procured from either Lions Eye Institute for Transplant and Research (Florida, USA) or Miracles in Sights (North Carolina, USA), with informed consent from the next of kin. All research performed with human derived tissue was carried out in accordance to the principles outlined in the Declaration of Helsinki. All corneo-scleral donor tissues were preserved and transported in Optisol-GS (Bausch & Lomb, New York, USA) at 4 °C until they were processed.