{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,6]],"date-time":"2026-07-06T20:01:57Z","timestamp":1783368117372,"version":"3.54.6"},"reference-count":50,"publisher":"MDPI AG","issue":"15","license":[{"start":{"date-parts":[[2022,8,3]],"date-time":"2022-08-03T00:00:00Z","timestamp":1659484800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"EU and the Hungarian Government","award":["EFOP-3.6.1-16-2016-00008"],"award-info":[{"award-number":["EFOP-3.6.1-16-2016-00008"]}]},{"name":"EU and the Hungarian Government","award":["SZTE \u00c1OK-KKA No 2018\/T\u00f3th-Moln\u00e1rE"],"award-info":[{"award-number":["SZTE \u00c1OK-KKA No 2018\/T\u00f3th-Moln\u00e1rE"]}]},{"name":"Albert Szent-Gy\u00f6rgyi Medical School, University of Szeged","award":["EFOP-3.6.1-16-2016-00008"],"award-info":[{"award-number":["EFOP-3.6.1-16-2016-00008"]}]},{"name":"Albert Szent-Gy\u00f6rgyi Medical School, University of Szeged","award":["SZTE \u00c1OK-KKA No 2018\/T\u00f3th-Moln\u00e1rE"],"award-info":[{"award-number":["SZTE \u00c1OK-KKA No 2018\/T\u00f3th-Moln\u00e1rE"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>Tear samples are considered in recent publications as easily, noninvasively collectible information sources for precision medicine. Their complex composition may aid the identification of biomarkers and the monitoring of the effectiveness of treatments for the eye and systemic diseases. Sample collection and processing are key steps in any analytical method, especially if subtle personal differences need to be detected. In this work, we evaluate the usability of a novel sample collection technique for human tear samples using phenol red threads (cotton thread treated with the pH indicator phenol red), which are efficiently used to measure tear volume in clinical diagnosis. The low invasiveness and low discomfort to the patients have already been demonstrated, but their applicability for proteomic sample collection has not yet been compared to other methods. We have shown, using various statistical approaches, the qualitative and quantitative differences in proteomic samples collected with this novel and two traditional methods using either glass capillaries or Schirmer\u2019s paper strips. In all parameters studied, the phenol red threads proved to be equally or even more suitable than traditional methods. Based on detectability using different sampling methods, we have classified proteins in tear samples.<\/jats:p>","DOI":"10.3390\/ijms23158647","type":"journal-article","created":{"date-parts":[[2022,8,3]],"date-time":"2022-08-03T20:52:01Z","timestamp":1659559921000},"page":"8647","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":6,"title":["An Extensive Study of Phenol Red Thread as a Novel Non-Invasive Tear Sampling Technique for Proteomics Studies: Comparison with Two Commonly Used Methods"],"prefix":"10.3390","volume":"23","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-5584-6869","authenticated-orcid":false,"given":"G\u00e1bor","family":"Kecskem\u00e9ti","sequence":"first","affiliation":[{"name":"Department of Medical Chemistry, Albert Szent-Gy\u00f6rgyi Medical School, University of Szeged, D\u00f3m t\u00e9r 8, H-6720 Szeged, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Edit","family":"T\u00f3th-Moln\u00e1r","sequence":"additional","affiliation":[{"name":"Department of Ophtalmology, Albert Szent-Gy\u00f6rgyi Health Centre, University of Szeged, Kor\u00e1nyi Fasor 10-11, H-6720 Szeged, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6466-8283","authenticated-orcid":false,"given":"Tam\u00e1s","family":"Jan\u00e1ky","sequence":"additional","affiliation":[{"name":"Department of Medical Chemistry, Albert Szent-Gy\u00f6rgyi Medical School, University of Szeged, D\u00f3m t\u00e9r 8, H-6720 Szeged, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8278-8038","authenticated-orcid":false,"given":"Zolt\u00e1n","family":"Szab\u00f3","sequence":"additional","affiliation":[{"name":"Department of Medical Chemistry, Albert Szent-Gy\u00f6rgyi Medical School, University of Szeged, D\u00f3m t\u00e9r 8, H-6720 Szeged, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2022,8,3]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"449","DOI":"10.1007\/s13167-021-00265-y","article-title":"Mass Spectrometry Analysis of Human Tear Fluid Biomarkers Specific for Ocular and Systemic Diseases in the Context of 3P Medicine","volume":"12","author":"Zhan","year":"2021","journal-title":"EPMA J."},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Ponzini, E., Santambrogio, C., De Palma, A., Mauri, P., Tavazzi, S., and Grandori, R. (2021). Mass Spectrometry-based Tear Proteomics for Noninvasive Biomarker Discovery. Mass Spectrom. Rev., 1\u201319.","DOI":"10.1002\/mas.21691"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"4876","DOI":"10.1021\/pr2004874","article-title":"Characterization of the Human Tear Metabolome by LC-MS\/MS","volume":"10","author":"Chen","year":"2011","journal-title":"J. Proteome Res."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"527","DOI":"10.1016\/j.preteyeres.2012.06.002","article-title":"Tear Analysis in Ocular Surface Diseases","volume":"31","author":"Zhou","year":"2012","journal-title":"Prog. Retin. Eye Res."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"189","DOI":"10.1080\/14789450.2017.1285703","article-title":"The Power of Tears: How Tear Proteomics Research Could Revolutionize the Clinic","volume":"14","author":"Zhou","year":"2017","journal-title":"Expert Rev. Proteom."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"83","DOI":"10.3892\/ijmm.2021.4916","article-title":"Critical Role of Mass Spectrometry Proteomics in Tear Biomarker Discovery for Multifactorial Ocular Diseases (Review)","volume":"47","author":"Ma","year":"2021","journal-title":"Int. J. Mol. Med."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"521","DOI":"10.1177\/0004563217695843","article-title":"Evaluation of Commonly Used Tear Sampling Methods and Their Relevance in Subsequent Biochemical Analysis","volume":"54","author":"Rentka","year":"2017","journal-title":"Ann. Clin. Biochem."},{"key":"ref_8","first-page":"390","article-title":"A Modified Schirmer Test: The Fine-Thread Method for Measuring Lacrimation","volume":"14","author":"Kurihashi","year":"1977","journal-title":"J. Pediatr. Ophthalmol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"23","DOI":"10.1167\/tvst.9.10.23","article-title":"Quantifying Sample Collection and Processing Impacts on Fiber-Based Tear Fluid Chemical Analysis","volume":"9","author":"Barmada","year":"2020","journal-title":"Transl. Vis. Sci. Technol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"16","DOI":"10.1167\/tvst.9.3.16","article-title":"Comparison of Capillary and Schirmer Strip Tear Fluid Sampling Methods Using Swath-Ms Proteomics Approach","volume":"9","author":"Aapola","year":"2020","journal-title":"Transl. Vis. Sci. Technol."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"40","DOI":"10.4103\/2229-516X.81979","article-title":"Evaluation of Phenol Red Thread Test versus Schirmer Test in Dry Eyes: A Comparative Study","volume":"1","author":"Vashisht","year":"2011","journal-title":"Int. J. Appl. Basic Med. Res."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"012017","DOI":"10.1088\/1757-899X\/336\/1\/012017","article-title":"Integration K-Means Clustering Method and Elbow Method for Identification of the Best Customer Profile Cluster","volume":"336","author":"Syakur","year":"2018","journal-title":"IOP Conf. Ser. Mater. Sci. Eng."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1700394","DOI":"10.1002\/pmic.201700394","article-title":"The Human Eye Proteome Project: Updates on an Emerging Proteome","volume":"18","author":"Ahmad","year":"2018","journal-title":"Proteomics"},{"key":"ref_14","unstructured":"(2018). Anterior Eye. Contact Lens Practice, Elsevier."},{"key":"ref_15","unstructured":"Ocran, E. (2022, June 25). Lacrimal Gland. Available online: https:\/\/www.kenhub.com\/en\/library\/anatomy\/lacrimal-gland."},{"key":"ref_16","first-page":"264","article-title":"Determination of Tear Volume and Tear Flow","volume":"5","author":"Mishima","year":"1966","journal-title":"Investig. Ophthalmol."},{"key":"ref_17","unstructured":"(2020). Tear Film\u2014Physiology and Disturbances in Various Diseases and Disorders. Ocular Surface Diseases\u2014Some Current Date on Tear Film Problem and Keratoconic Diagnosis, IntechOpen."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"60","DOI":"10.1016\/S1542-0124(12)70296-3","article-title":"Basal, Reflex, and Psycho-Emotional Tears","volume":"7","author":"Murube","year":"2009","journal-title":"Ocul. Surf."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"366","DOI":"10.1016\/j.jtos.2017.03.006","article-title":"TFOS DEWS II Tear Film Report","volume":"15","author":"Willcox","year":"2017","journal-title":"Ocul. Surf."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"2245","DOI":"10.1177\/1120672121998922","article-title":"Tear Fluid Collection Methods: Review of Current Techniques","volume":"31","author":"Szulc","year":"2021","journal-title":"Eur. J. Ophthalmol."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"5309","DOI":"10.1007\/s00216-016-9624-7","article-title":"Threads for Tear Film Collection and Support in Quantitative Amino Acid Analysis","volume":"408","author":"Avilov","year":"2016","journal-title":"Anal. Bioanal. Chem."},{"key":"ref_22","first-page":"456","article-title":"Investigation of the Human Tear Film Proteome Using Multiple Proteomic Approaches","volume":"14","author":"Nichols","year":"2008","journal-title":"Mol. Vis."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"137","DOI":"10.1016\/j.aanat.2012.10.001","article-title":"Schirmer Strip vs. Capillary Tube Method: Non-Invasive Methods of Obtaining Proteins from Tear Fluid","volume":"195","author":"Posa","year":"2013","journal-title":"Ann. Anat."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"1512","DOI":"10.1167\/iovs.11-7627","article-title":"Correlation of Tear Inflammatory Cytokines and Matrix Metalloproteinases with Four Dry Eye Diagnostic Tests","volume":"53","author":"Vandermeid","year":"2012","journal-title":"Investig. Ophthalmol. Vis. Sci."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"267","DOI":"10.1016\/0014-4835(76)90054-3","article-title":"The Origin of Some Enzymes in Tear Fluid, Determined by Comparative Investigation with Two Collection Methods","volume":"22","author":"Glasius","year":"1976","journal-title":"Exp. Eye Res."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"1853","DOI":"10.1002\/elps.200900755","article-title":"Post-Translation Modification of Proteins in Tears","volume":"31","author":"You","year":"2010","journal-title":"Electrophoresis"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"425","DOI":"10.1111\/vop.12876","article-title":"Influence of Schirmer Strip Wetness on Volume Absorbed, Volume Recovered, and Total Protein Content in Canine Tears","volume":"24","author":"Bertram","year":"2021","journal-title":"Vet. Ophthalmol."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"913","DOI":"10.1038\/sj.eye.6702052","article-title":"Phenol Red Thread Test vs Schirmer\u2019s Test: A Comparative Study","volume":"20","author":"Saleh","year":"2006","journal-title":"Eye"},{"key":"ref_29","first-page":"374","article-title":"The Effect of Collection Technique on Tear Composition","volume":"25","author":"Stuchell","year":"1984","journal-title":"Investig. Ophthalmol. Vis. Sci."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"135","DOI":"10.1016\/0014-4835(90)90223-H","article-title":"Effects of the Schirmer Test on the Fibrinolytic System in the Tear Fluid","volume":"50","author":"Stephens","year":"1990","journal-title":"Exp. Eye Res."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"152","DOI":"10.1097\/00006324-199302000-00011","article-title":"Schirmer Test. I. A Review","volume":"70","author":"Cho","year":"1993","journal-title":"Optom. Vis. Sci."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"127","DOI":"10.1097\/ICL.0000000000000025","article-title":"Comparative Study of Repeatability of Phenol Red Thread Test Versus Schirmer Test in Normal Adults in Saudi Arabia","volume":"40","author":"Masmali","year":"2014","journal-title":"Eye Contact Lens Sci. Clin. Pract."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"482","DOI":"10.1111\/j.1475-1313.2007.00500.x","article-title":"The Phenol Red Thread Test for Lacrimal Volume\u2014Does It Matter If the Eyes Are Open or Closed?","volume":"27","author":"Doughty","year":"2007","journal-title":"Ophthalmic Physiol. Opt."},{"key":"ref_34","first-page":"3130","article-title":"Water-Soluble Antioxidants in Human Tears: Effect of the Collection Method","volume":"42","author":"Choy","year":"2001","journal-title":"Investig. Ophthalmol. Vis. Sci."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1016\/j.ab.2015.04.002","article-title":"Single Unit Filter-Aided Method for Fast Proteomic Analysis of Tear Fluid","volume":"480","author":"Aass","year":"2015","journal-title":"Anal. Biochem."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"47","DOI":"10.1016\/j.clae.2017.09.012","article-title":"The Potential Influence of Schirmer Strip Variables on Dry Eye Disease Characterisation, and on Tear Collection and Analysis","volume":"41","author":"Mann","year":"2018","journal-title":"Contact Lens Anterior Eye"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"142","DOI":"10.1097\/00006324-200103000-00005","article-title":"What Does the Phenol Red Thread Test Actually Measure?","volume":"78","author":"Tomlinson","year":"2001","journal-title":"Optom. Vis. Sci."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"25","DOI":"10.1647\/2008-050","article-title":"Evaluation of the Phenol Red Thread Tear Test in Falconiformes","volume":"29","author":"Smith","year":"2015","journal-title":"J. Avian Med. Surg."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"436","DOI":"10.1111\/vop.12024","article-title":"Results of Phenol Red Thread Test in Clinically Normal Syrian Hamsters (Mesocricetus Auratus)","volume":"16","author":"Rajaei","year":"2013","journal-title":"Vet. Ophthalmol."},{"key":"ref_40","doi-asserted-by":"crossref","unstructured":"Akkurt Arslan, M., Kolman, I., Pionneau, C., Chardonnet, S., Magny, R., Baudouin, C., Brignole-Baudouin, F., and Kessal, K. (2022). Proteomic Analysis of Tears and Conjunctival Cells Collected with Schirmer Strips Using Timstof pro: Preanalytical Considerations. Metabolites, 12.","DOI":"10.3390\/metabo12010002"},{"key":"ref_41","doi-asserted-by":"crossref","unstructured":"Kall\u00f3, G., Emri, M., Varga, Z., Ujhelyi, B., Tozs\u00e9r, J., Csutak, A., and Csosz, \u00c9. (2016). Changes in the Chemical Barrier Composition of Tears in Alzheimer\u2019s Disease Reveal Potential Tear Diagnostic Biomarkers. PLoS ONE, 11.","DOI":"10.1371\/journal.pone.0158000"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"13363","DOI":"10.1038\/s41598-017-13817-y","article-title":"Proteomic Analysis of Human Lacrimal and Tear Fluid in Dry Eye Disease","volume":"7","author":"Jung","year":"2017","journal-title":"Sci. Rep."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"40","DOI":"10.1007\/s11095-019-2569-8","article-title":"Review of Biomarkers in Ocular Matrices: Challenges and Opportunities","volume":"36","author":"Tamhane","year":"2019","journal-title":"Pharm. Res."},{"key":"ref_44","first-page":"1549","article-title":"Alterations in Tear Biochemistry Associated with Postanesthetic Chronic Dry Eye Syndrome","volume":"81","author":"Zernii","year":"2016","journal-title":"Biochemistry"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"15","DOI":"10.1186\/s13167-016-0065-3","article-title":"Tear Fluid Biomarkers in Ocular and Systemic Disease: Potential Use for Predictive, Preventive and Personalised Medicine","volume":"7","author":"Hagan","year":"2016","journal-title":"EPMA J."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"10350","DOI":"10.1021\/acs.analchem.8b02172","article-title":"Surfactant Cocktail-Aided Extraction\/Precipitation\/On-Pellet Digestion Strategy Enables Efficient and Reproducible Sample Preparation for Large-Scale Quantitative Proteomics","volume":"90","author":"Shen","year":"2018","journal-title":"Anal. Chem."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"41","DOI":"10.1038\/s41592-019-0638-x","article-title":"DIA-NN: Neural Networks and Interference Correction Enable Deep Proteome Coverage in High Throughput","volume":"17","author":"Demichev","year":"2020","journal-title":"Nat. Methods"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"731","DOI":"10.1038\/nmeth.3901","article-title":"The Perseus Computational Platform for Comprehensive Analysis of (Prote)Omics Data","volume":"13","author":"Tyanova","year":"2016","journal-title":"Nat. Methods"},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"12648","DOI":"10.1038\/s41598-018-31154-6","article-title":"Instant Clue: A Software Suite for Interactive Data Visualization and Analysis","volume":"8","author":"Nolte","year":"2018","journal-title":"Sci. Rep."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"25","DOI":"10.1038\/75556","article-title":"Gene Ontology: Tool for the Unification of Biology","volume":"25","author":"Ashburner","year":"2000","journal-title":"Nat. Genet."}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/23\/15\/8647\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T00:03:46Z","timestamp":1760141026000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/23\/15\/8647"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2022,8,3]]},"references-count":50,"journal-issue":{"issue":"15","published-online":{"date-parts":[[2022,8]]}},"alternative-id":["ijms23158647"],"URL":"https:\/\/doi.org\/10.3390\/ijms23158647","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2022,8,3]]}}}