Optimization and evaluation of tear protein elution from Schirmer's strips in dry eye disease

• Published in: Indian journal of ophthalmology Vol. 71; no. 4; pp. 1413 - 1419
• Main Authors: Koduri, Madhuri, Prasad, Deeksha, Pingali, Tejaswini, Singh, Vijay, Shanbhag, Swapna, Basu, Sayan, Singh, Vivek
• Format: Journal Article
• Language: English
• Published: India Wolters Kluwer India Pvt. Ltd 01.04.2023; Medknow Publications and Media Pvt. Ltd; Medknow Publications & Media Pvt. Ltd; Wolters Kluwer - Medknow; Wolters Kluwer Medknow Publications
• Subjects: Bicarbonates; Composition; Diagnosis; dry eye disease; Dry eye syndromes; Dry Eye Syndromes - diagnosis; Dry Eye Syndromes - metabolism; Extraction (Chemistry); Eye; Eye diseases; Eye Proteins - metabolism; Health aspects; Humans; Immune response; Innate immunity; Mass spectroscopy; Medical examination; Methods; Octoxynol - metabolism; Original; Proteins; Proteolysis; schirmer's strip; stevens–johnson syndrome; tear proteomics; Tearing; Tears - metabolism; Wound healing; India; Stevens-Johnson syndrome; Dry eye disease; Schirmer's strip; tear proteomics; Stevens–Johnson syndrome
• ISSN: 0301-4738; 1998-3689
• DOI: 10.4103/IJO.IJO_2774_22

Purpose: Extraction of tear protein from Schirmer's strip is a prerequisite for the proper identification and screening of biomarkers in dry eye disease. The study compares different methods of extraction of tear proteins from the Schirmer's strip. Methods: Reflex tear was collected from healthy controls (HC; n = 12), Stevens-Johnson syndrome (SJS; n = 3) and dry eye disease (DED; n = 3) patients using capillary tube. This tear was used to measure the volume absorbed by Schirmer's strip per microliter. Different buffers (6) were used to compare the protein yield from the Schirmer's strip in four different conditions. The tear proteins extracted using the highest protein yield buffer were analyzed by mass spectrometry. Results: A linear relationship between the tear volume and wetting length was observed (r = 0.0.997, n = 6). The highest yield was observed after incubation of the Schirmer's strip in 100 mM ammonium bicarbonate (ABC) with 0.25% Nonidet P-40(NP-40) at 4°C for an hour (P < 0.00005). The in-solution digestion of tear eluted in the above condition 100 Mm ABC + 0.25% NP-40 with one-hour incubation yielded a total of 2119 proteins in HC, SJS, and DED. The unique protein observed in SJS and DED was 0.6% and 17.9%, respectively. The significantly expressed proteins are associated with innate immune response, proteolysis, wound healing, and defense response. Conclusion: A method for extraction of protein from Schirmer's strip was optimized for increase in protein yield from the tear sample. SJS and DED tear samples have unique protein signature. The study will aid in better design of tear protein-based experimental study.