Economic evaluation of endothelial keratoplasty techniques and penetrating keratoplasty in the Netherlands.

PURPOSE: To evaluate cost-effectiveness of penetrating keratoplasty (PK), femtosecond laser-assisted Descemet stripping endothelial keratoplasty (FS-DSEK), and Descemet stripping automated endothelial keratoplasty (DSAEK). DESIGN: Cost-effectiveness analysis based on data from a randomized multicenter clinical trial and a noncomparative prospective study. METHODS: Data of 118 patients with corneal endothelial dysfunction were analyzed in the economic evaluation. Forty patients were included in the PK group, 36 in the FS-DSEK group, and 42 in the DSAEK group. The primary incremental cost-effectiveness ratio (ICER) was the incremental costs per clinically improved patient, defined as a patient with a combined effectiveness of both a clinically improved BSCVA (defined as an improvement of at least 2 lines) and a clinically acceptable refractive astigmatism (defined as less than or equal to 3.0 diopters). Analysis was based on a 1-year follow-up period after transplantation. RESULTS: The percentage of treated patients who met the combined effectiveness measures was 52% for DSAEK, 44% for PK, and 43% for FS-DSEK. Mean total costs per patient were €6674 (US$7942), €12 443 (US$14 807), and €7072 (US$8416) in the PK group, FS-DSEK group, and DSAEK group, respectively. FS-DSEK was less effective and more costly compared to both DSAEK and PK. DSAEK was more costly but also more effective compared to PK, resulting in incremental costs of €4975 (US$5920) per additional clinically improved patient. CONCLUSIONS: The results of this study show that FS-DSEK was not cost-effective compared to PK and DSAEK. DSAEK, on the other hand, was more costly but also more effective compared to PK. Including societal costs, a longer follow-up period and preparation of the lamellar transplant buttons in a national cornea bank could improve the cost-effectiveness of DSAEK.

Quality of vision after femtosecond laser-assisted descemet stripping endothelial keratoplasty and penetrating keratoplasty: a randomized, multicenter clinical trial.

PURPOSE: To compare the quality of vision (straylight and contrast sensitivity) after femtosecond laser-assisted Descemet stripping endothelial keratoplasty (FS DSEK) and penetrating keratoplasty (PK). DESIGN: Prospective, randomized clinical trial. METHODS: setting: Multicenter (5 ophthalmic centers in The Netherlands). study population: Eighty eyes of 80 patients with corneal endothelial dysfunction were included and were randomized to FS DSEK or PK. observation procedures: FS DSEK and PK. main outcome measures: Straylight, contrast sensitivity, astigmatism, uncorrected visual acuity, best spectacle-corrected visual acuity (BSCVA), and visual symptom score. RESULTS: Straylight at 12 months was 1.37 ± 0.2 logarithm of straylight for FS DSEK and 1.46 ± 0.2 logarithm of straylight for PK (P = .151). During 12 months of follow-up, there was a significant improvement of straylight and contrast sensitivity after FS DSEK (P < .001) and PK (P < .001). The change of straylight and contrast sensitivity correlated significantly with the change of BSCVA after FS DSEK (r = -0.645; r = 0.580) and PK (r = -0.370; r = 0.659). The visual symptom score was comparable between the 2 groups during the 12 months of follow-up. CONCLUSIONS: Improvement of straylight and contrast sensitivity was significantly correlated with an improvement of BSCVA. Straylight and contrast sensitivity were improved significantly after FS DSEK and were comparable with those after PK, although BSCVA was slightly better in the PK group.

Economic evaluation of deep anterior lamellar keratoplasty versus penetrating keratoplasty in The Netherlands.

PURPOSE: To evaluate the cost effectiveness of deep anterior lamellar keratoplasty (DALK) versus penetrating keratoplasty (PK) in The Netherlands. DESIGN: Cost-effectiveness analysis alongside a randomized, multicenter clinical trial. METHODS: Fifty-three patients with corneal stromal pathologic features not affecting the endothelium were included with 28 patients in the DALK group and 25 in the PK group. Quality of life was measured before surgery and 3, 6, and 12 months after surgery. The main outcome measures were incremental cost-effectiveness ratios per clinically improved patient on the 25-item National Eye Institute Visual Functioning Questionnaire and per patient with endothelial cell loss of maximally 20% within the first year. RESULTS: Mean total bootstrapped costs per patient were €7607 (US$10,498) in the DALK group and €6552 (US$9042) in the PK group. The incremental cost-effectiveness ratios were €9977 (US$13,768) per clinically improved patient on the 25-item National Eye Institute Visual Functioning Questionnaire and €6900 (US$9522) per patient with cell loss of maximally 20%. In patients without perforation of the Descemet membrane, the incremental cost-effectiveness ratio was €5250 (US$7245) per patient. CONCLUSIONS: This study shows that DALK is more costly and more effective as compared with PK. Results on the 25-item National Eye Institute Visual Functioning Questionnaire were in favor of DALK, and endothelial cell loss in DALK patients remained stable after 6 months, whereas cell loss in PK patients continued. Furthermore, DALK procedures performed without perforation of the Descemet membrane were more effective. However, because it is unknown what society is willing to pay for an additional improved patient, cost effectiveness of DALK within a limited follow-up period of 12 months is unclear. Cost effectiveness of DALK may improve over time because of lower graft failure.

Endothelial cell loss and visual outcome of deep anterior lamellar keratoplasty versus penetrating keratoplasty: a randomized multicenter clinical trial.

OBJECTIVE: To compare endothelial cell (EC) loss, visual and refractive outcomes, and complications after deep anterior lamellar keratoplasty (DALK) and penetrating keratoplasty (PK). DESIGN: Randomized, multicenter clinical trial. PARTICIPANTS: Fifty-six eyes of 56 patients with a corneal stromal pathology not affecting the endothelium were randomized to DALK or PK. METHODS: The DALK procedure was performed according to Anwar's big-bubble technique. Patients underwent an ophthalmic examination preoperatively and 3, 6, and 12 months postoperatively. MAIN OUTCOME MEASURES: Endothelial cell loss, refractive and topographic astigmatism, spherical equivalent, uncorrected visual acuity, and best spectacle-corrected visual acuity (BSCVA) were measured, and complications were recorded. RESULTS: Endothelial cell loss was significantly higher after PK compared with DALK procedures performed without perforation of Descemet's membrane (12 months: 27.7% ± 11.1% vs. 12.9% ± 17.6%). The BSCVA was significantly better in the PK group at 3 and 6 months after surgery but was not significantly different 12 months after surgery (0.39 ± 0.3 logarithm of the minimum angle of resolution [logMAR] in DALK and 0.31 ± 0.3 logMAR in PK). At 12 months postoperatively, refractive and topographic astigmatism in the DALK and PK groups were -3.37 ± 2.3 diopters (D) and -3.76 ± 2.1 D (P = 0.53), and 3.57 ± 2.3 D and 4.16 ± 2.0 D (P = 0.34), respectively. (Micro)perforation of the Descemet's membrane occurred in 32% (9/28) of the DALK eyes, and 18% (5/28) of the patients required conversion to PK. Endothelial cell loss was not significantly different between DALK and PK when cases with perforation of Descemet's membrane were included in the (intention-to-treat) analysis (12 months: 19.1 ± 21.6 vs. 27.7 ± 11.1 P = 0.112). Rejection episodes were reported in 1 patient in the DALK group (epithelial rejection) and 3 patients in the PK group (all endothelial rejections). No graft failure occurred. CONCLUSIONS: One year after DALK performed without perforation of Descemet's membrane, EC loss is significantly lower, whereas the BSCVA is comparable to that in the PK group. In addition, no endothelial rejection occurred in the DALK group. However, Descemet's membrane perforation remains a major complication in DALK and warrants improvements to standardize the big-bubble technique.

Validation of tissue quality parameters for donor corneas, designated for emergency cases: corneal graft survival.

PURPOSE: To validate tissue quality parameters for donor corneas designated for emergency grafting for corneal graft survival. METHODS: In a longitudinal cohort follow-up study, 131 emergency penetrating grafts were studied. Grafts were performed with a pool of organ-cultured donor corneas designated for emergency grafting and prepared for immediate use with all safety tests performed. Assignation criteria were: corneas with a small superficial stromal opacity but meeting all selection criteria for PKP tissue and corneas without stromal opacity, but an endothelial cell density from 1800 to 2300 cells/mm(2) or mild polymegathism or pleomorphism. Cox multivariate regression analysis, Kaplan-Meier survival and log rank test were applied. RESULTS: Of the 131 keratoplasties, 115 could be followed. One eye was lost during surgery because of an expulsive bleeding. In 15 cases, a conjunctival transplantation finished off the penetrating graft. Corneal graft survival was not significantly related to the presence of PKP quality of the donor endothelium, neither with a cloudy graft nor with endothelial decompensation as the cause of failure. Main risk factors for a failed graft were vascularization of the host cornea (p=0.0001), the presence of a systemic auto immune disease in the recipient (p=0.003) and the disease leading to the (imminent) perforation and emergency graft (p=0.021). CONCLUSION: A selected pool of donor corneas designated for emergency grafting that does not interfere with the scheduled procedures allows more efficient and safe use of donor tissue in case of a(n) (imminent) perforation. Corneal graft survival rates justify the criteria for selection.

High subretinal fluid procoagulant activity in rhegmatogenous retinal detachment.

PURPOSE: An increased mRNA expression of genes related to blood coagulation has been demonstrated in an experimental retinal detachment model but has not yet been confirmed in human clinical specimens. Tissue factor (TF), the initiating factor of blood coagulation, may be a determinant of the extent of tissue injury after rhegmatogenous retinal detachment (RRD). This study was conducted to determine whether subretinal fluid and vitreous fluid collected from patients with RRD have a procoagulant effect. METHODS: Calibrated thrombin generation (CAT) was used to investigate the thrombogenic properties of 28 subretinal fluids collected during scleral buckling surgery for RRD. Further, the thrombogenic properties of vitreous fluids from RRD (n = 12), macular pucker (n = 5), macular hole (n = 6), and proliferative diabetic retinopathy (n = 5) were compared with the properties of eye bank eyes (n = 11), which served as control specimens. The procoagulant activity of TF was determined with Western blot analysis. RESULTS: The addition of subretinal fluid from all RRD patients (28/28, 100%) induced thrombin generation in normal and severely factor (F)XII-deficient plasma. Contrary to the subretinal fluid, the addition of vitreous fluids from various ocular disorders evoked very little thrombin generation in normal and severely FXII-deficient plasma (4/12, 33% RRD; 1/5, 20% macular pucker; 0/6, 0% macular hole; 0/5, 0% proliferative diabetic retinopathy; and 2/11, 18% eye bank eyes). The procoagulant activity in subretinal fluid was almost completely neutralized by antibodies against human TF. The presence of TF in subretinal fluid was confirmed by Western blot. CONCLUSIONS: Subretinal fluid of patients with RRD induces high procoagulant activity, determined by measuring the level of tissue factor.

Validation of tissue quality parameters for donor corneas designated for emergency use in preservation of the globe.

PURPOSE: To validate tissue quality parameters for donor corneas designated for emergency grafting to preserve the globe. METHODS: In a longitudinal cohort follow-up study, 151 emergency grafts in the Netherlands were studied. Grafts were performed with a pool of organ-cultured donor corneas designated for emergency grafting and prepared for immediate use with all safety tests performed. Assignation criteria were corneas with a small superficial stromal opacity but meeting all selection criteria for penetrating keratoplasty tissue and corneas without stromal opacity but an endothelial cell density from 1800 to 2300 cells per square millimeter or mild polymegethism or pleomorphism. Cox multivariate regression analysis, Kaplan-Meier survival, and log rank test were applied. RESULTS: All requests for corneal tissue were honored within 24 hours. Ninety-one patients showed a complete and 60 an imminent perforation. One hundred thirty-one penetrating grafts and 20 lamellar grafts have been used. The globes were saved in 140 of the 151 patients (92.7%). Globe preservation was not significantly related to the absence of penetrating keratoplasty quality of the donor endothelium, the type of grafting, the degree of vascularization in the host cornea, or diabetic disease in the recipient. The main risk factor for globe preservation was the presence of a systemic autoimmune disease in the recipient (P = 0.0021). CONCLUSIONS: A selected pool of donor corneas designated for emergency grafting, that does not interfere with the scheduled procedures, allows more efficient and safe use of donor tissue in case of an imminent perforation. Globe preservation rates justify the quality criteria for designation of this tissue.

Efficacy and safety of femtosecond laser-assisted corneal endothelial keratoplasty: a randomized multicenter clinical trial.

BACKGROUND: To evaluate the efficacy and safety of femtosecond laser-assisted endothelial keratoplasty (FLEK) versus penetrating keratoplasty (PK) in patients with corneal endothelial disease. METHODS: A randomized multicenter clinical trial of 80 eyes of 80 patients with corneal endothelial disease were randomized to FLEK or PK. Clinical outcomes (astigmatism and visual acuity) and incidence of postoperative complications were compared between the two groups. RESULTS: At 12 months, the percentage of eyes with a refractive astigmatism less than or equal to 3 diopters was higher in the FLEK group in comparison with the PK group (86.2% vs. 51.3%, P=0.004). The mean postoperative best corrected visual acuity was 20/70+/-2 lines in the FLEK group and 20/44+/-2 lines in the PK group (P<0.001), but the gain in the best corrected visual acuity between the two groups was not significantly different. The endothelial cell loss in the FLEK and PK group was 65+/-12% and 23+/-15% (P<0.001). The most common postoperative complication in the FLEK group was graft dislocation (27.8%). Wound healing related problems occurred in six eyes (15%) in the PK group and in none of the FLEK eyes. CONCLUSIONS: FLEK effectively reduces postoperative astigmatism and results in an absence of wound healing related problems in patients with endothelial disease. However, visual acuity is lower as compared with conventional PK, and the high level of endothelial cell loss warrants a modification of the insertion technique of the endothelial graft.

European eye bank association.

BACKGROUND: The European Eye Bank Association (EEBA) is a technical-scientific organization for eye banks. Founded in 1989 with the simple objective of sharing information on eye banking, the Association is today the leading pan-national association in Europe dedicated to the advancement of eye banking and an authoritative reference point for eye banks which work according to quality standards. METHODS: The Association establishes and maintains an agreed set of medical and technical standards, promotes the collection of data on eye bank activities and processes, provides opportunities for the discussion of all aspects of eye banking practice, including eye donor selection and procurement, relevant research and development, education and training in eye banking, and maintains linkage with national and international corneal transplant communities and relevant bodies. RESULTS: The recent introduction of a more structured and focused committee, a permanent secretariat, the development of a website has enabled the Association to establish closer links and collaborative activities with key regulatory bodies and to provide a more constant exchange of clinical, scientific and technical ideas and best practice with fellow professionals by means of its annual meetings, the EEBA directory and website, and a regular newsletter. CONCLUSION: The EEBA is a scientific organization committed to defining minimum standards and to encouraging eye banks to maintain the highest possible standards for quality and safety. Through its annual meetings, and the collection and exchange of detailed information from member eye banks, the Association can rightly claim to speak with a confident and representative voice on eye banking in Europe.

Histologic evaluation of human posterior lamellar discs for femtosecond laser Descemet's stripping endothelial keratoplasty.

PURPOSE: To evaluate the histologic changes in corneal structure after femtosecond laser preparation of posterior lamellar discs, more specifically, the smoothness of the stromal bed and the accuracy of the predicted depth of the horizontal lamellar cut. MATERIALS AND METHODS: Nineteen human donor eyes unsuitable for transplantation were used. Femtosecond laser was used to prepare a horizontal lamellar cut in donor corneas at a depth of 400 microm. Transmission electron microscopy images were used to evaluate the changes in the corneal structure and to measure the damage zone. Scanning electron microscopy images were used to determine the relative depth of the horizontal lamellar cut, and the stromal bed was examined to determine the smoothness of the surface. RESULTS: Transmission electron microscopy images showed a mean damage zone of 6.8 +/- 3.1 microm, which consisted of irregularly oriented collagen fibrils and electron-dense granular material. The collagen lamellae, both anteriorly and posteriorly of the damaged zone, showed a regular parallel configuration. The relative depth of the horizontal lamellar cut as percentage of the total corneal thickness in the center and periphery was 70.4% +/- 4.5% and 55.6% +/- 5.9%. Scanning electron microscopy images of the stromal bed showed a relatively smooth surface. CONCLUSION: The femtosecond laser is effective to prepare a deep horizontal lamellar cut in a standardized method. The stromal bed is smooth and without extensive adjacent tissue damage. The is thinner in the center and thicker at the edges, which may produce a mild hyperopic shift after femtosecond laser-assisted Descemet's stripping endothelial keratoplasty.

Preliminary results of femtosecond laser-assisted descemet stripping endothelial keratoplasty.

OBJECTIVE: To evaluate the preliminary visual results of femtosecond laser-assisted Descemet stripping endothelial keratoplasty (FS-DSEK). METHODS: We prospectively analyzed results of 20 consecutive patients with Fuchs endothelial dystrophy or aphakic/pseudophakic bullous keratopathy who underwent FS-DSEK. Best spectacle-corrected visual acuity (BSCVA), refraction, corneal topography, and endothelial cell density were measured preoperatively and 3 and 6 months after FS-DSEK. Corneal thickness was measured using an optical coherence tomography technique. RESULTS: The average BSCVA of 11 eyes with normal visual potential significantly improved from 20/110 +/- 4 lines to 20/57 +/- 1 line at 6 months (P < .007). At 6 months, the mean (SD) hyperopic shift was 2.24 (2.3) diopters (D). Preoperative and 6 months postoperative refractive astigmatism were -0.75 (0.9) D and -1.58 (1.1) D (P = .01), but the topographic astigmatism did not change postoperatively (P = .95). Mean (SD) endothelial cell density at 6 months was 1368 (425) cells/mm(2). There was a persistent deswelling of the graft up to 3 months postoperatively. Complications included graft dislocations requiring repositioning (20%), pupillary block glaucoma (5%), epithelial ingrowth (5%), and primary graft failure (5%). CONCLUSIONS: Femtosecond laser-assisted Descemet stripping endothelial keratoplasty was effective in treating endothelial failure with minimal induced refractive astigmatism, limited improvement of BSCVA, and induction of a hyperopic shift. Endothelial cell count and dislocation rate were significant, which may be related to the surgical technique.

Prospective clinical evaluation of McCarey-Kaufman and organ culture cornea preservation media: 14-year follow-up.

PURPOSE: To compare the outcome of corneal grafts preserved in McCarey-Kaufman (MK) medium versus organ culture after penetrating keratoplasties. METHODS: Paired corneas were stored in McCarey medium for 2-44 hours (mean 21 hours) and in organ culture (OC) for 144 -240 hours (mean 192 hours). Penetrating keratoplasties were performed by 2 surgeons in 2 groups of patients with keratoconus, matched for age. Each pair was transplanted by the same surgeon using the same technique. Visual acuity, central corneal thickness, and central endothelial cell density were assessed at 166 +/- 7.8 months postoperatively. RESULTS: Nine pairs of patients were recovered for a long-term follow-up. The mean endothelial cell densities for the MK and OC groups were 611 +/- 155 and 683 +/- 168 cells/mm, respectively, which were not significantly different. A first rapid cell loss rate of 2.07% and 2.52% per month and a second slow of 0.78% and 0.69% per month were observed in the respective groups. Individual values of best corrected visual acuity were all the same (value 1.00) for both groups. Corneal thicknesses were, respectively, 571 +/- 52 and 540 +/- 35 microm and were significantly different (P = 0.013). CONCLUSIONS: After 14 years of penetrating keratoplasties performed with corneas stored in MK versus OC, no significant differences were observed in visual acuity, endothelial cell density, and cell loss. The observed thinner grafts after OC compared with MK could not be explained.

Femtosecond laser-assisted inverted mushroom keratoplasty.

PURPOSE: To evaluate best-corrected visual acuity (BCVA), refractive outcome, corneal topography, optical coherence tomography, and endothelial cell density 12 months after femtosecond laser-assisted inverted mushroom keratoplasty. METHODS: We performed a prospective study of a surgical case series of 5 patients undergoing femtosecond laser-assisted inverted mushroom keratoplasty for pseudophakic bullous keratopathy or pre-Descemet X-linked ichthyosis. The femtosecond laser was used to create a top-hat configuration in the donor cornea and recipient cornea. Laser parameters were as follows: energy, 4.0 (anterior inner vertical side cut and horizontal lamellar cut) and 7.0 microJ (posterior outer vertical side cut); spiral pattern with a firing rate of 15 kHz. The size of the anterior inner diameter was 7.4 mm in the donor cornea and 7.0 mm in the recipient cornea. The posterior outer diameter was 9.0 mm in all eyes. RESULTS: At 6 and 12 months after surgery, all corneal grafts were clear and showed an excellent adaptation of the lamellar donor and recipient wound surfaces. At 12 months postoperatively, BCVA averaged 20/32 (range, 20/60-20/20), refractive cylinder averaged -3.20 +/- 2.0 D, topographical cylinder averaged 3.26 +/- 2.1 D, and the mean endothelial cell density was 1793 +/- 491 cells/mm2 (range, 954-2237 cells/mm2). The mean central corneal thickness and thickness of the posterior shelf was 517 +/- 3 and 175 +/- 8 microm, respectively. CONCLUSIONS: The femtosecond laser-assisted inverted mushroom keratoplasty shows good promise in surgical treatment of corneal diseases. The multiplanar fit between the donor and recipient cornea allows early suture removal and visual rehabilitation.

Eye bank issues: II. Preservation techniques: warm versus cold storage.

Most of the tissue used for penetrating keratoplasty is issued through eye banks that store the corneoscleral button either in hypothermic storage at 2-6 degrees C or in organ culture at 31-37 degrees C. These two preservation techniques differ in technical aspects, tissue evaluation possibilities, storage time and microbiological safety. Hypothermic storage is simple and requires little expensive equipment. In general a pre-storage evaluation of the endothelium is performed by specular microscopy and storage time is usually around 7-10 days. Organ culture is a relatively complicated technique requiring more expertise and well-equipped facilities. Evaluation of the endothelium is not only performed before storage, but is routinely performed after storage through the use of light microscopy. With organ culture the allowed storage period is longer, up to four weeks. The vulnerability of organ culture to microbial contamination can be turned into an advantage because it allows the detection of residual micro-organisms on the cornea before surgery. Both preservation techniques seem to result in similar graft survival. The method of choice for preservation of the donor cornea is dictated by a number of factors mentioned in this review and this helps to explain the geographical differences in the use of the different techniques.

Corneal endothelial viability after femtosecond laser preparation of posterior lamellar discs for Descemet-stripping endothelial keratoplasty.

PURPOSE: To evaluate the feasibility of the femtosecond (FS) laser in preparation of posterior lamellar discs (PLDs) and to study the effect on endothelial cell (EC) viability for Descemet-stripping endothelial keratoplasty. METHODS: Fourteen human donor bulbi unsuitable for transplantation were used. A horizontal lamellar cut was prepared in the donor cornea with an FS laser by using a raster and/or a spiral spot pattern. The control group consisted of the paired cornea of the same donor. EC density was evaluated before and after preservation in organ culture. The PLD was stripped from the anterior part by using either a forceps or a blunt dissection technique. The damage to the endothelium was evaluated. RESULTS: EC loss after organ storage was not statistically significant between the FS cornea group and the control group in the 15- (7.7% +/- 6.9% and 8.9% +/- 8.1%, respectively; P = 0.78) and 30-kHz (4.3% +/- 4.0% and 3.7% +/- 3.6%, respectively; P = 0.75) group. There was no significant effect of laser frequency (15 vs. 30 kHz) on EC loss (7.7% vs. 4.3%, P = 0.25). Dissection by using a forceps stripping technique resulted in higher EC loss than that with a blunt dissection technique (13.0% vs. 6.5%, P = 0.03). CONCLUSIONS: EC loss after FS laser lamellar cutting is not dependent on the frequency (ie, energy level) of the laser. A blunt dissection technique of PLDs resulted in acceptable EC loss and supports the clinical use of the FS laser for the preparation of PLDs.

Femtosecond-laser-assisted Descemet's stripping endothelial keratoplasty.

To our knowledge, we describe the first patient with pseudophakic bullous keratoplasty treated with femtosecond-laser-assisted endothelial keratoplasty. A 5.5 mm corneoscleral tunnel incision was made; after Descemet's membrane was stripped, an 8.0 mm posterior lamellar corneal disk prepared with a femtosecond laser was inserted into the anterior chamber against the recipient cornea without the use of corneal sutures. Four months postoperatively, the posterior corneal disk was clear and the induced astigmatism was 2.1 diopters, demonstrating a functional corneal endothelial layer. The femtosecond laser offers a new surgical approach for minimally invasive endothelial keratoplasty in corneal endothelial disorders.

Biocompatibility of trypan blue with human corneal cells.

OBJECTIVE: To quantify the toxicity of trypan blue on human corneal cells according to exposure time and concentration. METHODS: Three in vitro experiments were performed. (1) We exposed cultured human corneal fibroblasts to trypan blue (0.0001% to 0.1%) in Eagle modified minimum essential medium (EMEM) or phosphate-buffered saline (PBS) for 15 minutes to 24 hours. Cytotoxicity was evaluated by Mosmann's colorimetric 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MMT) assay. (2) We exposed human corneas in EMEM for 24 hours to trypan blue (0.001% to 0.1%). Fellow donor corneas served as controls. Endothelial survival was evaluated morphologically and by cell density assessment. (3) We morphologically compared the endothelial viability of human donor corneas after exposure to 0.1% trypan blue for 5 to 30 minutes with control corneas. RESULTS: In experiment 1, trypan blue in EMEM was not significantly toxic at concentrations of 0.005% or lower. Higher concentrations were toxic only after exposure to trypan blue for at least 6 hours. In PBS, significant toxicity was found after exposure to 0.1% trypan blue for 30 minutes or longer. Lower concentrations were toxic after longer exposures. In experiment 2, exposure to 0.01% and 0.1% trypan blue for 24 hours resulted in significant loss in cell density. At lower concentrations, the endothelium was affected only morphologically. In experiment 3, endothelial morphology changed in control corneas and after exposure to 0.1% trypan blue for as little as 5 minutes. After 30-minute exposure, morphologic deterioration was more pronounced. CONCLUSIONS: Trypan blue was toxic in vitro to corneal endothelium and corneal fibroblasts at higher concentrations and notably longer exposure times. Toxicity was less in EMEM than in PBS. Clinical Relevance At commonly used concentrations, both during cataract surgery and in the cornea bank, trypan blue is safe for corneal cells. At higher concentrations or longer exposures, however, caution is warranted.