An Extensive Study of Phenol Red Thread as a Novel Non-Invasive Tear Sampling Technique for Proteomics Studies: Comparison with Two Commonly Used Methods

Tear samples are considered in recent publications as easily, noninvasively collectible information sources for precision medicine. Their complex composition may serve the identification of biomarkers and monitoring the effectiveness of treatments in eye and systematic 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 discomfort to the patients have already been demonstrated, but their applicability for proteomic sample collection was not yet 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’s 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. Data Independent Analysis (DIA) LC-MS experiments were perfomed on tear samples collected using different methods. A combined spectral library was created from Gas Phase Fractionation (GPF) DIA measurements of pool samples using DIA-NN 1.8. The spectral library is uploaded in TSV format. Quantitative data matrices exported by DIA-NN at proteingroup (pg) and precursor (pr) levelare also uploaded from the analysis of tearsamples from both eyes of 10 healthy individuals collected by the three sample collection methods.