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The plasma OPN concentration was measured using the enzyme-linked immunosorbent assay (ELISA) method with TiterZyme® Osteopontin detection ELISA kits (TiterZyme, USA). Plates were incubated with PBS-Tween containing 1% fetal calf serum at 37°C for 60 min to block nonspecific binding. Subsequently, OPN standards and serum samples were added to the plate. After incubation at 37°C for 60 min, 100 μl of antibody was added to each well with the exception of the blank well and the plates were incubated at room temperature for 60 min. The plates were washed with PBS, 100 μl of conjugate was added to each well with the exception of the blank well, and the plates were incubated at room temperature for 30 min. The plates were washed and the substrate solution and color were added and measured at 570 and 590 nm by a microtiter plate reader (Model 550, BIO-RAD, Hercules, California). The OPN concentration was calculated using linear regression by plotting the average net OD for each standard versus the human OPN concentration in each standard. An approximation of a straight line was made through the points and the concentration of human OPN in the serum was determined by interpolation.. per years was significant at (p=0.05).. endeavour has been supported by methodologies such as data mining,.

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Among patients diagnosed with type A influenza, laninamivir was prescribed to significantly older patients (typically aged ≥ 10 years), whereas oseltamivir was prescribed to significantly younger patients (aged < 10 years) since the 2011‐2012 season (P < .05) (Figure 2 B‐1, 2). As seen in Figure 2C, only around 20 patients were diagnosed with type B influenza during each season. Zanamivir was prescribed to approximately 20% of patients with type B influenza. This NAI was preferred for use in patients with type B influenza compared to those with type A.. In addition to oral manifestations and rheumatological implications, Sjӧgren's syndrome seems to play a role in TMD especially as regards to muscular contracture (high prevalence of myalgia to muscle palpation and positive endfeel) and joint disease (greater pop-noise and deflection during mandibular opening), causing an increase in orofacial pain and an altered chewing function.

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