Balanced salt solution bss

The AH samples of DME eyes were collected at baseline, and at 1, 3 and 12 months post first SMPL application, while the sampling of the non-diabetic control group was performed only once, as no changes of proteins’ concentration were expected through the time. All patients underwent the standard preoperative procedure: peri-ocular skin disinfection with povidone-iodine 10% (ESO-JOD, ECOLAB, Agrate Brianza, Italy), instillation of sterile lidocaine 4% (Alpha Intes, Napoli, Italy), irrigation of conjunctival sac with povidone-iodine 5% (Oftasteril, Alpha Intes) for two minutes and washing of the eye with Balanced Salt Solution (BSS; Alcon, Fort Worth, TX, USA). From the anterior chamber, AH (150–200 μL) was aspirated with an insulin syringe (31-gauge needle). AH samples were then collected in a single vial containing 10 μL of a cocktail of protease inhibitors (Pierce Biotechnology, Rock-ford, IL, USA) and quickly stored at −70 °C until analysis. The total protein content was quantified with a digital spectrophotometer (NanoDrop; Thermo Fisher Scientific Inc., Waltham, MA, USA) and protein concentrations were calculated by means of the linearized standard curve (BSA) and the A280 software. Thereafter, AH samples underwent sonication (VibraCell; Sonics, Newton, CT, USA) and centrifugation to collect clear supernatant (13000 rpm/7 min).

Cysteamine hydrochloride BioXtra, cysteamine purity 98%, cystamine impurity standard, orthophosphoric acid 85% and acid 1-heptanosulphonic, sodium salt were obtained from Sigma-Aldrich (St. Louis, MO, USA). Hyaluronic acid, with a molecular weight of 10⁶ Da and a purity of 97%, was purchased from Acofarma (Madrid, Spain). Ethylenediaminetetraacetic acid disodium salt (EDTA) was obtained from Fagron (Barcelona, Spain). HPLC grade acetonitrile was purchased from Thermo Fisher Scientific (Frederick, MA, USA). Balanced salt solution (BSS) was purchased from Alcon (Geneva, Switzerland). Hydrochloric acid 1M was obtained from Carlo Erba (Barcelona, Spain). Ultrapure water MilliQ, Merck Millipore (Madrid, Spain) was used. Class I 15 mL amber glass dropper bottles were used as primary packaging (Fagron, Barcelona, Spain).