Development and Validation of a Multiplex Electrochemiluminescence Immunoassay to Evaluate Dry Eye Disease in Rat Tear Fluids

Overview

Journal Sci Rep

Specialty Science

Date 2023 Jul 27

PMID 37500810

Authors

Agnese Compagnone

An Matheeussen

Linda De Vooght

Paul Cos

Affiliations

Soon will be listed here.

Abstract

Dry eye disease (DED) is a challenge in ophthalmology. Rat models represent valuable tools to study the pathophysiology and to develop novel treatments. A major challenge in DED research is detecting multiple biomarkers in a low tear volume sample. Multiplex immunoassays for DED rat research are missing. We have developed a multiplex electrochemiluminescence immunoassay (ECLIA) to detect three biomarkers for DED: MMP-9, IL-17 and ICAM-1. Tears, used as matrix, were collected from six healthy Wistar rats. Assays were run based on the U-Plex Meso Scale Diagnostics (MSD) platform, by two independent operators according to the EMA guideline on bioanalytical method validation. Linear mixed, regression models were fit to perform the statistical analysis on the range of concentrations for the chosen analytes. During optimization, it has observed that incubation time, temperature and agitation affected the robustness of the protocol. ECLIA optimum conditions include the use of antibodies at 0.5 µg/ml concentration and 1 h incubation at room temperature with shaking. Precision met the acceptance criteria in the chosen range: 1062-133 pg/ml for ICAM-1, 275-34.4 pg/ml for IL-17, 1750-219 pg/ml for MMP-9. Accuracy and linearity were acceptable for a broader range. This is the first report of a validated ECLIA that allows measurements of three relevant DED biomarkers in rat tear fluids.

References

Zaleska-Zmijewska A, Strzemecka E, Wawrzyniak Z, Szaflik J . Extracellular MMP-9-Based Assessment of Ocular Surface Inflammation in Patients with Primary Open-Angle Glaucoma. J Ophthalmol. 2019; 2019:1240537. PMC: 6470422. DOI: 10.1155/2019/1240537. View

Garbutcheon-Singh K, Carnt N, Pattamatta U, Samarawickrama C, White A, Calder V . A Review of the Cytokine IL-17 in Ocular Surface and Corneal Disease. Curr Eye Res. 2018; 44(1):1-10. DOI: 10.1080/02713683.2018.1519834. View

Auerswald H, Eng C, Lay S, In S, Eng S, Vo H . Rapid Generation of In-House Serological Assays Is Comparable to Commercial Kits Critical for Early Response to Pandemics: A Case With SARS-CoV-2. Front Med (Lausanne). 2022; 9:864972. PMC: 9121123. DOI: 10.3389/fmed.2022.864972. View

Stapleton F, Alves M, Bunya V, Jalbert I, Lekhanont K, Malet F . TFOS DEWS II Epidemiology Report. Ocul Surf. 2017; 15(3):334-365. DOI: 10.1016/j.jtos.2017.05.003. View

Zheng X, de Paiva C, Li D, Farley W, Pflugfelder S . Desiccating stress promotion of Th17 differentiation by ocular surface tissues through a dendritic cell-mediated pathway. Invest Ophthalmol Vis Sci. 2010; 51(6):3083-91. PMC: 2891467. DOI: 10.1167/iovs.09-3838. View

Stern M, Schaumburg C, Pflugfelder S . Dry eye as a mucosal autoimmune disease. Int Rev Immunol. 2013; 32(1):19-41. PMC: 3587314. DOI: 10.3109/08830185.2012.748052. View

Wu X, Chen X, Ma Y, Lin X, Yu X, He S . Analysis of tear inflammatory molecules and clinical correlations in evaporative dry eye disease caused by meibomian gland dysfunction. Int Ophthalmol. 2020; 40(11):3049-3058. PMC: 7550292. DOI: 10.1007/s10792-020-01489-z. View

Gao J, Morgan G, Tieu D, Schwalb T, Luo J, Wheeler L . ICAM-1 expression predisposes ocular tissues to immune-based inflammation in dry eye patients and Sjögrens syndrome-like MRL/lpr mice. Exp Eye Res. 2004; 78(4):823-35. DOI: 10.1016/j.exer.2003.10.024. View

Kunert K, Tisdale A, Stern M, Smith J, Gipson I . Analysis of topical cyclosporine treatment of patients with dry eye syndrome: effect on conjunctival lymphocytes. Arch Ophthalmol. 2000; 118(11):1489-96. DOI: 10.1001/archopht.118.11.1489. View

10.

Stern M, Pflugfelder S . What We Have Learned From Animal Models of Dry Eye. Int Ophthalmol Clin. 2017; 57(2):109-118. PMC: 5347474. DOI: 10.1097/IIO.0000000000000169. View

11.

Joossen C, Baan A, Moreno-Cinos C, Joossens J, Cools N, Lanckacker E . A novel serine protease inhibitor as potential treatment for dry eye syndrome and ocular inflammation. Sci Rep. 2020; 10(1):17268. PMC: 7560718. DOI: 10.1038/s41598-020-74159-w. View

12.

Andreasson U, Perret-Liaudet A, van Waalwijk van Doorn L, Blennow K, Chiasserini D, Engelborghs S . A Practical Guide to Immunoassay Method Validation. Front Neurol. 2015; 6:179. PMC: 4541289. DOI: 10.3389/fneur.2015.00179. View

13.

Jie Y, Xu L, Wu Y, Jonas J . Prevalence of dry eye among adult Chinese in the Beijing Eye Study. Eye (Lond). 2008; 23(3):688-93. DOI: 10.1038/sj.eye.6703101. View

14.

Tsubota K, Fujihara T, Saito K, Takeuchi T . Conjunctival epithelium expression of HLA-DR in dry eye patients. Ophthalmologica. 1998; 213(1):16-9. DOI: 10.1159/000027387. View

15.

Chang L, Zhao J, Guo F, Ji H, Zhang L, Jiang X . Comparative Evaluation and Measure of Accuracy of ELISAs, CLIAs, and ECLIAs for the Detection of HIV Infection among Blood Donors in China. Can J Infect Dis Med Microbiol. 2020; 2020:2164685. PMC: 7443234. DOI: 10.1155/2020/2164685. View

16.

Lian Y, Fu H, Xu X, Ju C . Application Effect and Accuracy Analysis of Electrochemiluminescence Immunoassay and Enzyme-Linked Immunosorbent Assay in the Serological Test of Hepatitis B Virus. Evid Based Complement Alternat Med. 2022; 2022:9371497. PMC: 9371851. DOI: 10.1155/2022/9371497. View

17.

Perez V, Pflugfelder S, Zhang S, Shojaei A, Haque R . Lifitegrast, a Novel Integrin Antagonist for Treatment of Dry Eye Disease. Ocul Surf. 2016; 14(2):207-15. DOI: 10.1016/j.jtos.2016.01.001. View

18.

Scarpellini C, Ramos Llorca A, Lanthier C, Klejborowska G, Augustyns K . The Potential Role of Regulated Cell Death in Dry Eye Diseases and Ocular Surface Dysfunction. Int J Mol Sci. 2023; 24(1). PMC: 9820812. DOI: 10.3390/ijms24010731. View

19.

Oh H, Kim C, Lee J, Yang J . Effects of Quercetin in a Mouse Model of Experimental Dry Eye. Cornea. 2015; 34(9):1130-6. DOI: 10.1097/ICO.0000000000000543. View

20.

Kaku N, Nishimura F, Shigeishi Y, Tachiki R, Sakai H, Sasaki D . Performance of anti-SARS-CoV-2 antibody testing in asymptomatic or mild COVID-19 patients: A retrospective study in outbreak on a cruise ship. PLoS One. 2021; 16(9):e0257452. PMC: 8478248. DOI: 10.1371/journal.pone.0257452. View