Abstract
Introduction. To evaluate clinical feasibility and reproducibility of cytometric bead assay (CBA) in nondiluted vitreous samples of patients with age-related macular degeneration (ARMD), diabetic macular edema (DME), and central retinal vein occlusion (CRVO). Methods. Twelve patients from a single clinics day qualified for intravitreal injections (ARMD n = 6, DME n = 3, CRVO n = 3) and underwent a combination treatment including a single-site 23 gauge core vitrectomy which yielded a volume of 0.6 mL undiluted vitreous per patient. Interleukin-6 (IL-6), vascular endothelial growth factor isoform A (VEGF-A), and monocyte chemo-attractant protein-1 (MCP-1) were assessed directly from 0.3 mL at the same day (fresh samples). To assess the reproducibility 0.3 ml were frozen for 60 days at −80°, on which the CBA was repeated (frozen samples). Results. In the fresh samples IL-6 was highest in CRVO (median IL-6 55.8 pg/mL) > DME (50.6) > ARMD (3.1). Highest VEGF was measured in CRVO (447.4) > DME (3.9) > ARMD (2.0). MCP-1 was highest in CRVO (595.7) > AMD (530.8) > DME (178). The CBA reproducibility after frozen storage was examined to be most accurate for MCP1 (P = 0.91) > VEGF (P = 0.68) > IL-6 (P = 0.49). Conclusions. CBA is an innovative, fast determining, and reliable technology to analyze proteins in fluids, like the undiluted vitreous, which is important to better understand ocular pathophysiology and pharmacology. There is no influence of intermittent storage at −80° for the reproducibility of the CBA.
Highlights
To evaluate clinical feasibility and reproducibility of cytometric bead assay (CBA) in nondiluted vitreous samples of patients with age-related macular degeneration (ARMD), diabetic macular edema (DME), and central retinal vein occlusion (CRVO)
Twelve patients were included in the present study: six age-related macular degeneration (ARMD) patients with an active occult choroidal neovascularization and an average central macular thickness of 368 ± 45 μm measured by spectral optical coherence tomography (OCT; 3D OCT-2000; Topcon Deutschland, Willich, Germany), three patients with diabetic macular edema (DME; age 66.8 ± 10.5 years) and a mean central macular thickness of 508 ± 108 μm, and three patients (69.5 ± 14.2) with central retinal vein occlusion (CRVO) with a history of 11 weeks ± 3 weeks of CRVO and a central macular thickness of 557 ± 125 μm
In the fresh samples IL-6 was highest in CRVO > DME (50.6; 3.5–89) > ARMD (3.1; 2.4–60.2)
Summary
To evaluate clinical feasibility and reproducibility of cytometric bead assay (CBA) in nondiluted vitreous samples of patients with age-related macular degeneration (ARMD), diabetic macular edema (DME), and central retinal vein occlusion (CRVO). Maier et al were the only ones to demonstrate that vitreous and serum concentrations of cytokines, including vascular endothelial growth factor (VEGF), determined by CBA showed a strong correlation with those measured by ELISA in patients with advanced diabetic retinopathy that had to undergo threeport vitrectomy [6]. It is not clear, from a laboratory point of view, if the CBA results are reproducible after intermittent storage at −80◦C, which often happens in the clinical and laboratory routine. It is interesting to assess the CBA technique for the detection of inflammatory and proangiogenic cytokines in patients that would
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