Trypan Blue free DMEK: Use of Brilliant Blue combined with Lutein / Zeaxanthin for staining of Descemet’s membrane during DMEK surgery

Abstract

Purpose: Trypan blue, known for its toxic effect on corneal endothelial cells, is the standard dye used for visualization of Descemet's membrane (DM) in Descemet membrane endothelial keratoplasty (DMEK). It was previously shown that for lens capsular visualization less corneal endothelial cells become apoptic when brilliant blue (BB) is used instead of trypan blue.The purpose of this study was to investigate the efficacy and safety of a dye, based on lutein/zeaxanthin (L/Z) and BB, in staining corneal endothelium and DM for DMEK.

Methods: In vitro efficacy was determined in 60 cadaveric eyes by filling the anterior chamber with different combinations of L/Z and BB (0.01-20% and 0.0125-0.025%, respectively). These formulations were in contact with corneal endothelial cells for 30 seconds, then removed by mechanical aspiration and evaluated for their staining capacity.<br /> In vitro safety experiments with the same formulations were performed in human corneal endothelial cells (HCE) and their cellular toxicity was assessed with WST-1 colorimetric assay. Statistical significance was determined by one-way factorial repeated measurements analysis of variance (ANOVA).<br /> DMEK surgery was performed as previously described using L/Z 1% combined with 0.025 % BB (Retidyne Cap^®) for staining of DM during graft preparation in 5 patients

Results: Contact between L/Z combined with BB and the intraocular structures resulted in a greenish staining of the corneal endothelium and anterior capsule. The intensity of staining correlated with the concentration of BB (highest in 0.025 % BB). No significant differences were observed in the cytotoxic profiles after the exposure to each dye solution at all time periods.<br /> Graft preparation and transplantation was successful in all 5 DMEK surgeries. Staining of DM was faint at the beginning but became more intense during the following staining steps during the preparation.

Conclusions: A dye solution containing lutein and zeaxanthin combined with brilliant blue efficiently stained the corneal endothelium in human cadaveric eyes. These solutions showed a safe profile in an HCE model.<br /> Moreover, the combination of 1 % L/Z and 0.025 % BB was suitable for graft preparation and transplantation in DMEK. L/Z and BB are promising and less toxic alternatives to trypan blue for the use in DMEK.