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M. Esmaeelpour, M. Boulton, J. Cai, P.J. Murphy; The Effect of Tear Collection Technique on the Accuracy of Total Tear Protein, Major Proteins, and Serum Albumin . Invest. Ophthalmol. Vis. Sci. 2005;46(13):4398.
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Purpose: To compare the accuracy of two different tear collection techniques by monitoring the concentrations of standard bovine serum albumin (BSA) and total tear protein. To determine the effect of collection method on the major tear proteins: IgA, lactoferrin (LF), lysozyme (LZ) and serum albumin (SA). To investigate their volume absorbance efficiency. Methods: Seven subjects were recruited (3 female, 4 male; age = 19–31ys). Two non–stimulated tear samples were taken, one with a glass capillary tube (CT) and one with a cellulose rod (CR), on three different days. Each collection was completed before noon and limited to a maximum duration of 3min, with 1/2h between samples. To extract the tears, the cellulose rod was centrifuged at 15000g/3min/4°C, and the tears expelled from the tubes. All samples were assayed with the Bradford test for their total protein content. In a second study, 27 subjects (17 female, 10 male; age = 22–35ys) were recruited. One tear sample was taken with each technique over a maximum time of 1min 45sec. Samples were run subjected to onSDS–PAGE and visualizedstained by with silver stain. For the major tear protein study, 16 subjects (7 female, 9 male; age = 22–31ys) were recruited. Tear samples were taken similar to the second study but gels were stained by blue stain. Commercial standards used were human SA (0.25g/l), IgA (1.8g/l), LF (2.5g/l), and egg LZ (2.5g/l). Results: The average tear protein concentrations found were: CR 12.9g/l ±2; CT 11.9g/l ±2.8. The average tear volumes collected were: CR 7.6µl ±7.4; CT 2.7µl ±1.3. The BSA concentration measured for the CR was increased from the actual inserted concentration, due to volume changes from absorption and evaporation effects. The concentration was reduced for the CT. This produced a significant difference between the two techniques (p<0.001). However, for the tear protein concentration no significant difference was shown (p=0.17), because of higher volumes and variation in subjects. In the second study, 22 subjects had at least 0.5µl of tears collected with both techniques. For all subjects there was no clinical significant increase to prove serum leakage. Results for major tear proteins were obtained from 12 subjects. No significant difference was shown between both collection methods for IgA (p=0.45), LF (p=0.22) and LZ (p=0.29) although significantly higher volumes were collected with the CR (p=0.004). Conclusions: The cellulose rod offers a suitable alternative to the standard glass capillary tube.
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