Red and green defocus curves and duochrome test in different age groups / Curvas de desenfoque rojo y verde y prueba de duocromo en diferentes grupos de edad

Purpose: To compare the eye defocus curves (DCs) obtained with stimuli on red, green, and white backgrounds and to investigate the applicability of the duochrome test (DT) in different age groups. Methods: 12 elderly (ELD: 59.3 ± 3.9 years) and 8 young (YG: 22.1 ± 1.1 years) subjects were recruited. An optometric assessment with the DT was carried out to obtain the subjective refraction at distance. DCs at distance on green, white, and red backgrounds were measured and the following parameters were deduced: dioptric difference between red-green, green-white, red-white focal positions (minima of the DCs), best corrected visual acuity (BCVA), and widths of the DCs for red, green, and white. Results: The DC difference between the green-white focal positions (mean ± standard deviation) was -0.12±0.17 diopters (D) (ELD, p = 0.012) and -0.11±0.12 D (YG, p = 0.039), while the red-white difference was not statistically significant. The DC red-green difference was 0.20±0.16 D (ELD, p = 0.002) and 0.18±0.18 D (YG, p = 0.008). The ELD BCVA with green background was significantly worse than BCVA with red (p = 0.007) and white (p = 0.007). The mean value of the DC's width in ELD for green (1.01±0.36 D) was higher than for red (0.77±0.21 D) and for white (0.84±0.35 D), but with no statistical significance. Conclusion: Both age groups showed a slight focusing preference for red when using white light. Moreover, ELD showed a worse BCVA with a green compared to a red background. Despite these results deduced by DC analyses, these aspects do not compromise the possibility of using the DT in clinical practice both in the young and in the elderly. Furthermore, the difference of about 0.20 D between red-green DC in both groups confirms the clinical appropriateness of the widespread use of 0.25 D step as the standard minimum difference in power between correcting lenses.(AU)

In-vitro dehydration kinetics coefficient of Kalifilcon A and other contact lens materials.

In contact lens (CL) wear, dehydration needs to be tailored to avoid dryness and related symptoms. In this view, this work aims to assess and compare the in-vitro dehydration kinetics of five CL materials, including the newly developed Kalifilcon A CL. At 36 °C and 60% relative humidity, the in-vitro dehydration kinetics of the different CLs were compared using a gravimetric method. CLs were analyzed either after a rinse of a few seconds in preservative-free saline solution or after a 24-h incubation in the same solution. A model based on the Fick diffusion equation was employed to deduce a water kinetics coefficient, providing insights into water diffusion within the polymeric matrix. The study reveals that all materials exhibit a non-Fickian dehydration behavior, with significant differences in dehydration kinetics coefficients and dehydration rate slopes. Etafilcon A and Omafilcon A, both hydrogel CLs, exhibit a similar behavior, different compared to the pattern shown by Senofilcon A and Delefilcon A, silicone-hydrogel CLs. Notably, Kalifilcon A, despite being a silicone-hydrogel, displays a hydration behavior reminiscent of hydrogel CLs.

Comparing automated and manual assessments of tear break-up time using different non-invasive devices and a fluorescein procedure.

To evaluate the agreement and repeatability of an automated topography-based method for non-invasive break-up time (NIBUT) analyses in comparison with two other NIBUT procedures, the fluorescein procedure (fBUT), and with the manual assessment with the same device. In the first experiment, a semi-randomised crossover study was performed on forty-three participants (23.1 ± 2.1 years). NIBUT measurements were collected in a randomised order, in both eyes of participants with EasyTear View + (Easytear, Rovereto), Polaris, and Sirius + (CSO, Firenze). Then a fBUT was collected. The overall measurement procedure was repeated in a further session (retest) on the same day. In a second experiment, a retrospective randomised crossover study was performed on eighty-five NIBUT videos previously recorded by the Sirius+. Two observers assessed manually the videos and the NIBUTs were compared with the automatic ones. In the first experiment, ANOVA showed a significant difference between the four measures in both eyes (p < 0.001). Significant differences were found in the paired comparisons between each NIBUT procedure and fBUT (Wicoxon; p < 0.05). Sirius+ resulted in agreement only with Polaris in the left eye. Correlations between all NIBUT procedures resulted in statistical significance in both eyes. All procedures showed very good test-rest reliability. In the second experiment, a significant correlation between automated and manual NIBUT was found, but also a significant statistical difference between the two measurements, although clinically negligible (0.3 s). The investigated NIBUT devices perform differently from each other (and from fBUT), so they cannot be considered interchangeable. The automated measure of NIBUT with Sirius+ has a negligible clinical difference compared to manual assessment on the same device.

Red and green defocus curves and duochrome test in different age groups.

PURPOSE: To compare the eye defocus curves (DCs) obtained with stimuli on red, green, and white backgrounds and to investigate the applicability of the duochrome test (DT) in different age groups. METHODS: 12 elderly (ELD: 59.3 ± 3.9 years) and 8 young (YG: 22.1 ± 1.1 years) subjects were recruited. An optometric assessment with the DT was carried out to obtain the subjective refraction at distance. DCs at distance on green, white, and red backgrounds were measured and the following parameters were deduced: dioptric difference between red-green, green-white, red-white focal positions (minima of the DCs), best corrected visual acuity (BCVA), and widths of the DCs for red, green, and white. RESULTS: The DC difference between the green-white focal positions (mean ± standard deviation) was -0.12±0.17 diopters (D) (ELD, p = 0.012) and -0.11±0.12 D (YG, p = 0.039), while the red-white difference was not statistically significant. The DC red-green difference was 0.20±0.16 D (ELD, p = 0.002) and 0.18±0.18 D (YG, p = 0.008). The ELD BCVA with green background was significantly worse than BCVA with red (p = 0.007) and white (p = 0.007). The mean value of the DC's width in ELD for green (1.01±0.36 D) was higher than for red (0.77±0.21 D) and for white (0.84±0.35 D), but with no statistical significance. CONCLUSION: Both age groups showed a slight focusing preference for red when using white light. Moreover, ELD showed a worse BCVA with a green compared to a red background. Despite these results deduced by DC analyses, these aspects do not compromise the possibility of using the DT in clinical practice both in the young and in the elderly. Furthermore, the difference of about 0.20 D between red-green DC in both groups confirms the clinical appropriateness of the widespread use of 0.25 D step as the standard minimum difference in power between correcting lenses.

Modulation of Alpha-Synuclein Conformational Ensemble and Aggregation Pathways by Dopamine and Related Molecules.

Dopaminergic neurons are constantly threatened by the thin boundaries between functional α-synuclein (AS) structural disorder and pathogenic aggregation, and between dopamine (DA) neurotransmitter activity and accumulation of cytotoxic by-products. The possibilities of developing drugs for Parkinson's disease (PD) depend on our understanding of the molecular mechanisms that cause or accompany the pathological structural changes in AS. This review focuses on the three interconnected aspects of AS conformational transitions, its aggregation pathways and ligand binding. Specifically, the interactions of AS with DA, DA metabolites, DA analogs and DA agonists are considered. Recent advances in the field are discussed with reference to the structural properties of AS and the methodologies employed. Although several issues are still object of debate, salient structural features of the protein, the aggregates and the ligands can be identified, in the hope of fueling experimental and computational approaches to the discovery of novel disease-modifying agents.

Italian translation and validation of the Contact Lens Dry Eye Questionnaire-8 (CLDEQ-8).

PURPOSE: To translate and validate an Italian version of the CLDEQ-8 (CLDEQ-8_IT). METHODS: The study was carried out in two phases. In the first phase, a cross-cultural adaptation of CLDEQ-8 to Italian was performed by forward and backward translation in sequence. In the second phase, a multi-centre study was conducted for the validation of the questionnaire. Validity CLDEQ-8_IT was evaluated against three gestalt questions: overall opinion of soft contact lenses (CLs), global self-assessments of eye sensitivity and eye dryness. Reliability was evaluated by test-retest assessment in a subgroup of subjects. Finally, the psychometric properties of CLDEQ-8_IT were explored by Rasch analysis. RESULTS: Two hundred and forty soft CL wearers, fluent Italian speakers (73 males and 167 females), between 18 and 70 years of age were enrolled. A significant correlation was found between CLDEQ-8_IT and each of the three Gestalt questions. The cutoff score of 12 points demonstrated the best balance between sensitivity and specificity in differentiating wearers grading their CLs as "Excellent/Very good" from those reporting their overall opinion as "Good/Fair/Poor". The Intraclass Correlation Coefficient between test and retest was 0.88 (95% CI: 0.81-0.92). Finally, infit and outfit statistics using Rasch analysis for the 8 items were in a good range, however Principal Components Analysis revealed a certain degree of multi-dimensionality of the instrument. Also, item 8 analysis could be computed after merging the last two response categories. CONCLUSION: The CLDEQ-8_IT showed very good validity and reliability in measuring symptoms of CL wearers, comparable to the original English language version. A cut-off of 12 was confirmed as yielding the best balance between sensitivity and specificity in detecting CL wearers who could benefit from clinical management of their CL-related symptoms. Collapsing of the response options 5 and 6 in the last item of questionnaire could optimise its functioning.

Physical Properties and Interaction With the Ocular Surface of Water-Gradient Contact Lenses.

ABSTRACT: Since the introduction of silicone hydrogel contact lenses, many silicone-hydrogel materials have been produced, including water-gradient contact lenses with a silicone hydrogel core and a thin hydrogel outer layer (e.g., delefilcon A, verofilcon A, and lehfilcon A). Their properties have been investigated in various studies assessing both the chemical-physical characteristics and the comfort, but the overall picture is not always consistent. In this study, water-gradient technology is reviewed by looking at basic physical properties both in vitro and in vivo and at the interaction with the human ocular surface. Surface and bulk dehydration, surface wetting and dewetting, shear stress, interaction with tear components and with other environmental compounds, and comfort are discussed.

Contact Lens Wear Induces Alterations of Lactoferrin Functionality in Human Tears.

The tear film is a complex matrix composed of several molecular classes, from small metal ions to macromolecules. Contact lens (CL) wear can affect the protein homeostasis of the tear film, by accumulating deposits on the CL surface and/or altering their structural and functional properties. This work investigates the effect of CL wear on lactoferrin (Lf), one of the most abundant tear proteins, known as an unspecific biomarker of inflammation. Tears from eight volunteers were collected and analyzed after alternated periods of CL wear and without CL. The experimental approach is to probe Lf into unprocessed human tears by the peculiar fluorescence emission originating from complex formation of Lf with terbium (Tb3+) at the iron-binding sites. The experimental data indicate that CL wear does not significantly affect the total amount of Lf. On the other hand, Lf affinity for Tb3+ is reduced upon CL wear, suggesting relevant changes in Lf structure and possible alterations of protein functionality. Future studies based on this approach will help define CL features (material, lens-care solution, wearing time, etc.) with minimal effects on tear protein activity, in order to obtain more biocompatible and comfortable devices.

Native mass spectrometry for the investigation of protein structural (dis)order.

A central challenge in structural biology is represented by dynamic and heterogeneous systems, as typically represented by proteins in solution, with the extreme case of intrinsically disordered proteins (IDPs) [1-3]. These proteins lack a specific three-dimensional structure and have poorly organized secondary structure. For these reasons, they escape structural characterization by conventional biophysical methods. The investigation of these systems requires description of conformational ensembles, rather than of unique, defined structures or bundles of largely superimposable structures. Mass spectrometry (MS) has become a central tool in this field, offering a variety of complementary approaches to generate structural information on either folded or disordered proteins [4-6]. Two main categories of methods can be recognized. On one side, conformation-dependent reactions (such as cross-linking, covalent labeling, H/D exchange) are exploited to label molecules in solution, followed by the characterization of the labeling products by denaturing MS [7-11]. On the other side, non-denaturing ("native") MS can be used to directly explore the different conformational components in terms of geometry and structural compactness [12-16]. All these approaches have in common the capability to conjugate protein structure investigation with the peculiar analytical power of MS measurements, offering the possibility of assessing species distributions for folding and binding equilibria and the combination of both. These methods can be combined with characterization of noncovalent complexes [17, 18] and post-translational modifications [19-23]. This review focuses on the application of native MS to protein structure and dynamics investigation, with a general methodological section, followed by examples on specific proteins from our laboratory.

Viscoelastic properties of the human tear film.

When considering eye blinking from a tribological perspective, tear viscosity is expected to play a fundamental role. The application of rheological techniques to describe the tear film dates back to the late 1980s, but there has been a continuous need of reappraisal due to new findings in tear film biochemistry and to the development of new methods of physico-chemical characterization. This review provides an overview on tear rheological behavior by analyzing the peer-reviewed literature on this topic. Specifically, examples of in-vitro and in-vivo viscosity measurements are detailed, highlighting experimental criticalities and the need of a standard convention for rheological techniques to compare data across different studies, of analyses on tears of single individuals even within the limits of the low volume available, and of a further development of in-vivo techniques. Then, the controversial role of specific tear components on viscosity is discussed, together with the alterations associated to dry eye disease and contact lens wear. Finally, an updated focus is reported on the viscosity of artificial tears formulations.

Mass spectrometry-based tear proteomics for noninvasive biomarker discovery.

The lacrimal film has attracted increasing interest in the last decades as a potential source of biomarkers of physiopathological states, due to its accessibility, moderate complexity, and responsiveness to ocular and systemic diseases. High-performance liquid chromatography-mass spectrometry (LC-MS) has led to effective approaches to tear proteomics, despite the intrinsic limitations in sample amounts. This review focuses on the recent progress in strategy and technology, with an emphasis on the potential for personalized medicine. After an introduction on lacrimal-film composition, examples of applications to biomarker discovery are discussed, comparing approaches based on pooled-sample and single-tear analysis. Then, the most critical steps of the experimental pipeline, that is, tear collection, sample fractionation, and LC-MS implementation, are discussed with reference to proteome-coverage optimization. Advantages and challenges of the alternative procedures are highlighted. Despite the still limited number of studies, tear quantitative proteomics, including single-tear investigation, could offer unique contributions to the identification of low-invasiveness, sustained-accessibility biomarkers, and to the development of personalized approaches to therapy and diagnosis.

In vitro affinity for nicotine of soft contact lenses of different materials.

Smoking is a risk factor for the development of microbial keratitis and corneal infiltrates in contact lens (CL) wearers. It is still unknown if this risk is directly associated with the presence of nicotine in the eye and if adherence of nicotine on the CL can enhance these effects. A better understanding of the interaction between nicotine and CL materials could offer insights to explain this risk associated with smoking. The aim of this work was to compare the affinity of nicotine to different soft CL materials. CLs from FDA groups I, II, IV, and V were incubated in a 2-mM nicotine solution for 24 h and then in a 0.9% saline solution for the next 24 h. The amount of absorbed and released nicotine per CL was deduced as a function of time (t) by ultraviolet (UV) spectrophotometry and normalised to the mass of the hydrated CL. The data were described by the equation y = b -a t-1, where a and b are constants, and b represents the mass reached at the plateau after ~ 10 min of exposure. Groups IV and V displayed the highest (0.80 ± 0.09 µg) and lowest (0.27 ± 0.08 µg) nicotine absorption per mg of hydrated CL, respectively. The CL affinity for nicotine could be ascribed to the interaction between the positive charge of nicotine pyrrolidine nitrogen and the negative charges of the CLs, especially for the ionic IV group.

Tear-Based Vibrational Spectroscopy Applied to Amyotrophic Lateral Sclerosis.

Biofluid analysis by optical spectroscopy techniques is attracting considerable interest due to its potential to revolutionize diagnostics and precision medicine, particularly for neurodegenerative diseases. However, the lack of effective biomarkers combined with the unaccomplished identification of convenient biofluids has drastically hampered optical advancements in clinical diagnosis and monitoring of neurodegenerative disorders. Here, we show that vibrational spectroscopy applied to human tears opens a new route, offering a non-invasive, label-free identification of a devastating disease such as amyotrophic lateral sclerosis (ALS). Our proposed approach has been validated using two widespread techniques, namely, Fourier transform infrared (FTIR) and Raman microspectroscopies. In conjunction with multivariate analysis, this vibrational approach made it possible to discriminate between tears from ALS patients and healthy controls (HCs) with high specificity (∼97% and ∼100% for FTIR and Raman spectroscopy, respectively) and sensitivity (∼88% and ∼100% for FTIR and Raman spectroscopy, respectively). Additionally, the investigation of tears allowed us to disclose ALS spectroscopic markers related to protein and lipid alterations, as well as to a reduction of the phenylalanine level, in comparison with HCs. Our findings show that vibrational spectroscopy is a new potential ALS diagnostic approach and indicate that tears are a reliable and non-invasive source of ALS biomarkers.

Single-Tear Proteomics: A Feasible Approach to Precision Medicine.

Lacrimal fluid is an attractive source of noninvasive biomarkers, the main limitation being the small sample amounts typically collected. Advanced analytical methods to allow for proteomics profiling from a few microliters are needed to develop innovative biomarkers, with attractive perspectives of applications to precision medicine. This work describes an effective, analytical pipeline for single-tear analysis by ultrahigh-resolution, shotgun proteomics from 23 healthy human volunteers, leading to high-confidence identification of a total of 890 proteins. Highly reproducible quantification was achieved by either peak intensity, peak area, or spectral counting. Hierarchical clustering revealed a stratification of females vs. males that did not emerge from previous studies on pooled samples. Two subjects were monitored weekly over 3 weeks. The samples clustered by withdrawal time of day (morning vs. afternoon) but not by follow-up week, with elevated levels of components of the immune system in the morning samples. This study demonstrates feasibility of single-tear quantitative proteomics, envisaging contributions of this unconventional body fluid to individualized approaches in biomedicine.

The effect of active smoking, passive smoking, and e-cigarettes on the tear film: An updated comprehensive review.

Active tobacco smoking, passive smoking, and e-cigarette smoking have been associated with different systemic and ocular diseases. The precorneal tear film plays an important role in eye health and its analysis can provide useful information on ocular status. This review investigates the effects of different types of smoking on the precorneal tear film, by analyzing the peer-reviewed literature on this topic. Specifically, tear evaporation rate, stability, volume, ferning, osmolarity, and physical composition (lipids and proteins) of tear film are detailed. Most of the reported works show that cigarette smoking reduces tear film stability and quality by affecting its components. This review highlights that smoking severely affects the tear film, but a single test is not sufficient to determine these effects because smoking can impact different parts of the eye.

Profiling Dopamine-Induced Oxidized Proteoforms of ß-synuclein by Top-Down Mass Spectrometry.

The formation of multiple proteoforms by post-translational modifications (PTMs) enables a single protein to acquire distinct functional roles in its biological context. Oxidation of methionine residues (Met) is a common PTM, involved in physiological (e.g., signaling) and pathological (e.g., oxidative stress) states. This PTM typically maps at multiple protein sites, generating a heterogeneous population of proteoforms with specific biophysical and biochemical properties. The identification and quantitation of the variety of oxidized proteoforms originated under a given condition is required to assess the exact molecular nature of the species responsible for the process under investigation. In this work, the binding and oxidation of human ß-synuclein (BS) by dopamine (DA) has been explored. Native mass spectrometry (MS) has been employed to analyze the interaction of BS with DA. In a second step, top-down fragmentation of the intact protein from denaturing conditions has been performed to identify and quantify the distinct proteoforms generated by DA-induced oxidation. The analysis of isobaric proteoforms is approached by a combination of electron-transfer dissociation (ETD) at each extent of modification, quantitation of methionine-containing fragments and combinatorial analysis of the fragmentation products by multiple linear regression. This procedure represents a promising approach to systematic assessment of proteoforms variety and their relative abundance. The method can be adapted, in principle, to any protein containing any number of methionine residues, allowing for a full structural characterization of the protein oxidation states.

Lactoferrin Concentration in Human Tears and Ocular Diseases: A Meta-Analysis.

Purpose: To evaluate the potential of lactoferrin (Lf) as a diagnostic biomarker for ocular diseases using a meta-analytic approach. Methods: All original studies reporting an estimate of the average Lf concentration in healthy subjects and those affected by ocular diseases were searched up to March 2020. The DerSimonian and Laird method was used to calculate the random effects pooled mean difference and the corresponding 95% confidence interval (CI) in Lf concentration between healthy subjects and those affected by dry eye (DE), Sjögren syndrome (SS), and diabetic retinopathy, separately. The presence of between-study heterogeneity was evaluated using the Cochran's Q test and the I2 index. Stratified analyses were performed to assess potential sources of heterogeneity and influence and cumulative analyses to evaluate the robustness of the results obtained. Publication bias was also evaluated using funnel plot and the Egger's test. Results: The pooled mean differences in Lf concentrations between healthy subjects and those with DE, Sjögren syndrome, and diabetic retinopathy were respectively 0.62 (95% CI, 0.35-0.89) for DE, 3.78 (95% CI, -6.64 to 14.17), and 0.19 (95% CI, -4.00 to 4.39). Regarding DE, the stratified analysis showed that geographical area (P value Q test < 0.0001) and sample size (P < 0.0005) were sources of heterogeneity. Moreover, no study substantially influenced the results obtained and the pooled mean difference became statistically significant after a sample size of 220. Publication bias may affect the results of DE. Conclusions: The results of the current meta-analysis suggest that Lf level in tears is a good candidate as dry eye syndrome diagnostic biomarker.

Methionine oxidation in α-synuclein inhibits its propensity for ordered secondary structure.

α-Synuclein (AS) is an intrinsically disordered protein highly expressed in dopaminergic neurons. Its amyloid aggregates are the major component of Lewy bodies, a hallmark of Parkinson's disease (PD). AS is particularly exposed to oxidation of its methionine residues, both in vivo and in vitro Oxidative stress has been implicated in PD and oxidized α-synuclein has been shown to assemble into soluble, toxic oligomers, rather than amyloid fibrils. However, the structural effects of methionine oxidation are still poorly understood. In this work, oxidized AS was obtained by prolonged incubations with dopamine (DA) or epigallocatechin-3-gallate (EGCG), two inhibitors of AS aggregation, indicating that EGCG promotes the same final oxidation product as DA. The conformational transitions of the oxidized and non-oxidized protein were monitored by complementary biophysical techniques, including MS, ion mobility (IM), CD, and FTIR spectroscopy assays. Although the two variants displayed very similar structures under conditions that stabilize highly disordered or highly ordered states, differences emerged in the intermediate points of transitions induced by organic solvents, such as trifluoroethanol (TFE) and methanol (MeOH), indicating a lower propensity of the oxidized protein for forming either α- or ß-type secondary structures. Furthermore, oxidized AS displayed restricted secondary-structure transitions in response to dehydration and slightly amplified tertiary-structure transitions induced by ligand binding. This difference in susceptibility to induced folding could explain the loss of fibrillation potential observed for oxidized AS. Finally, site-specific oxidation kinetics point out a minor delay in Met-127 modification, likely due to the effects of AS intrinsic structure.

Structural characterization of aerogels derived from enzymatically oxidized galactomannans of fenugreek, sesbania and guar gums.

Aerogels are obtained by laccase/2,2,6,6-tetramethylpiperidin-1-yl)oxyl (TEMPO)-oxidation of galactomannans (GMs) from the leguminous plants fenugreek (Trigonella foenum-graecum), sesbania (Sesbania bispinosa) and guar (Cyamopsis tetragonolobus). GM oxidation in aqueous solutions causes a viscosity increase, resulting in structured and stable hydrogels. Upon lyophilization, water-insoluble aerogels are obtained, capable of water uptake up to several times their own weight. The materials derived from the three gums have been analyzed and compared. Chemical modifications have been studied by electrospray-ionization mass spectrometry (ESI-MS) and Fourier-transform infrared spectroscopy (FTIR). Polymer structure has been determined by liquid-state and semi-quantitative solid-state nuclear magnetic resonance (NMR) spectroscopy and chemical stability by incubation under variable conditions of pH, ionic strength and solvent nature. The results show that hydrogel formation is due to oxidation of primary hydroxyl groups to carbonyl and carboxyl groups and subsequent formation of hemiacetal and ester bonds. Fenugreek displays the highest stability, compared to guar and sesbania rehydrated aerogels. This feature could be interpreted by its higher degree of substitution (Gal:Man = 1:1) and consequently higher amount of galactose primary alcohols, leading to more extensive crosslinking.

Conformational Characterization and Classification of Intrinsically Disordered Proteins by Native Mass Spectrometry and Charge-State Distribution Analysis.

Intrinsically disordered proteins (IDPs) are systematically under-represented in structural proteomics studies. Their structural characterization implies description of the dynamic conformational ensembles populated by these polymers in solution, posing major challenges to biophysical methods. "Native" MS (native-MS) has emerged as a central tool in this field, conjugating the unique MS analytical power with structurally meaningful descriptors, like solvent-accessible surface area (SASA) and collisional cross section (CCS). This review summarizes recently published papers comparing native-MS and solution methods, with a focus on charge-state-distribution (CSD) analysis for IDP conformational analysis. The results point to substantial agreement, supporting structural interpretation of native-MS spectra of IDPs. The discussion is integrated with data from our group on "synthetic" IDPs, obtained by reduction and alkylation of natively folded proteins, whose fold is stabilized by disulfide bridges. Finally, an MS-based compaction index (CI) is introduced, evaluating SASA with reference to globular and fully disorder proteins. Such a parameter can be calculated for single conformers or the whole conformational ensemble, offering a continuous index for IDP comparison and classification.