Effect of tear fluid sampling and processing on total protein quantity and electrophoretic pattern.

Human tears contain more than 1500 proteins that could be diagnostically relevant. To date, numerous candidates on a biomarker of protein origin were identified for ocular and systemic diseases. However, the suitable sampling method is still the subject of discussion. To address the need for a description of sampling methods properties for possible clinical analyses, we studied a total protein concentration and electrophoretic pattern of tear fluid collected by capillary tubes, Schirmer strips, cellulose microsponges, and flushing. The total protein concentration was 4.339 µg/µL ± 1.905 µg/µL, 0.967 µg/µL ± 0.117 µg/µL, 0.022 µg/µL ± 0.016 µg/µL, and 0.008 µg/µL ± 0.006 µg/µ for the capillary tubes, Schirmer strips, flushing, and cellulose microsponges, respectively. Sodium dodecyl sulfate polyacrylamide electrophoresis showed the different patterns of tear proteins obtained by the above-mentioned sampling methods. These differences could originate from the use of a bigger amount of extraction reagent that was not used in the case of capillary tubes, and retention of the proteins by strips and sponges. Taken together, capillary tubes, Schirmer strips, cellulose microsponges, and flushing represent sensitive and convenient sampling methods for tear fluid collection. For the isolation of proteins from strips and sponges, and for the flushing, less than 100 µL of a reagent should be used to ensure the sufficient concentration of the biomarkers in a trace amount.

Effects of Humic Acids in Chronic Lead Poisoning.

Chronic exposure to lead causes disruption to energy production mechanisms and tissue damage, in particular through its binding to thiol groups and competition for zinc binding sites. We investigated the possibility of preventing the consequences of chronic lead poisoning by administration of three different doses of humic acids (HAs) into feed with the aim of establishing an effective HA dose. During the 10-week experiment, a sub-lethal dose of lead acetate was given to rats during the first 5 weeks, with continuous administration of HA over 10 weeks. Measurements were taken to determine the content of the metals Pb, Mn, Cu, Fe and Zn; the metalloid Se; and selected antioxidant markers in the heart, liver, kidney and plasma after the first, fifth and tenth weeks of experiment. The administration of lead and HAs clearly affects the redistribution of the elements and the activity of the antioxidant enzymes. This fact was particularly highlighted in the lead-only group as, within the experiment, significantly higher Pb concentrations were found only in the plasma of this group. However, in the group with 1% HA administered with lead, we observed a rise in Zn concentrations in the organs and the deposition of Fe into the liver. Decreased glutathione reductase activity in the plasma and balanced reduced glutathione concentrations indicated sufficient efficiency of redox reactions. SOD activities were among those affected most strongly, with only the 1% HA group showing no effect on heavy metal redistribution as a result of HA administration.

Noninvasive diagnostic methods for diabetes mellitus from tear fluid.

Diabetes mellitus and prolonged hyperglycemia can cause diabetic retinopathy. Diabetic retinopathy arises from damage to retinal vessels and, in its final stages, causes blindness. The early stages are often asymptomatic and although regular screening of diabetic patients is recommended, the beginning of diabetic retinopathy is insufficiently detected. The diagnostic potential of fluorescence spectroscopy, infrared spectroscopy and atomic force microscopy as the untraditional methods for diabetes mellitus was investigated using tear fluid. In our pilot study the structural changes of tear fluid of patients with diabetes mellitus after insulin and oral antidiabetic drug treatment was compared with healthy subjects. The results of analysis, infrared spectroscopy and atomic force microscopy confirmed structural changes in tear fluid of patients in comparison with the tear fluid of healthy subjects. Using new experimental laboratory methods in future could contribute to an improvement in diagnosis of diabetes mellitus and other selected ocular diseases using tear fluid.

Diagnostic potential of tears in ophthalmology.

In research circles, there is an increasing need to seek and identify new methods in the diagnosis of pathologies, monitoring the progression of the disease and response to treatment. The sensitivity of detection technologies has improved markedly, and enables the quantification of analyses in very small quantities. Tears represent a biological material with ever increasingly developing possibilities in the diagnosis of various pathologies. Our objective was to compile a basic overview of the diagnostic potential of tears via a summary of the potential lachrymal biomarkers of various pathologies. The article contains descriptions of protein biomarkers studied particularly in recent years, which correlate with a certain ocular pathology (dry eye, allergy, glaucoma etc.). It also summarises the results published to date in the field of systemic pathologies in patients with scleroderma, cystic fibrosis, diabetes mellitus, multiple sclerosis, cancers and Parkinson's disease. It concentrates on proteomic analyses, with the aim of improving the effectiveness of markers which could be used in future also in the timely diagnosis of ocular pathologies in clinical practice.Key words: Tears, proteins, biomarker, ocular pathologies, systemic pathologies.