Baseline Proteomic and Biochemical Characterization of Lens and Aqueous Humour Proteins in Native Sheep: Establishing Molecular References for Early Ocular Disease Detection

Abstract

Early diagnosis of ocular diseases in livestock is limited due to the lack of baseline molecular reference data for intraocular proteins. The  present study aimed to characterize biochemical and proteomic profiles of lens and aqueous humour proteins in healthy sheep and to  isolate major crystallin fractions from ovine lens tissues. Eyes from clinically healthy sheep (n = 10) were collected from a local abattoir,  and aqueous humour and lens tissues were processed for biochemical and electrophoretic analysis. Total protein concentration was  estimated using a colorimetric method, and protein profiling was performed using SDS-polyacrylamide gel electrophoresis (SDS-PAGE).  Lens proteins were further purified using DEAE anion-exchange chromatography with sodium chloride gradient elution. The mean  total protein concentration was significantly higher in lens homogenates (32.45 ± 1.28 mg/mL) compared with aqueous humour (1.86  ± 0.14 mg/mL), indicating the selective permeability of the blood-aqueous barrier. SDS-PAGE analysis revealed protein bands ranging  from 10-55 kDa in aqueous humour and dominant crystallin bands between 18-30 kDa in lens extracts. Chromatographic purification of  ovine lens proteins resulted in several fractions, with a prominent band observed at approximately 18.4 kDa in fractions eluted with 0.4  M NaCl. This band corresponded to gamma crystallin based on molecular weight estimation. The findings provide baseline biochemical  references for ocular proteins in sheep and demonstrate an effective approach for purification of crystallin proteins from ovine lenses.  These molecular insights may facilitate future biomarker discovery for early diagnosis of ocular diseases in livestock.