engleski

The first flow cytometric determination of salivary cell differential count

Human saliva is a complex exocrine secretion which contains proteins, hormones, antibodies, ions, bacteria and cells derived from salivary glands, cervical fluid, tonsils, the oral mucosa, bronchopulmonar and oropharyngeal secretions. Development of technologies for saliva-based diagnostics is in the beginning and little is known about its cellular components. We developed a method to quantify leukocytes in paraffin-stimulated whole human saliva (SWS) by flow cytometry and performed reliability study on 8 healthy volunteers (4 males, 4 females, mean age). SWS samples were taken every morning, three days in a row and analyzed immediately. Peripheral blood samples were taken only on the first day. After filtration of the samples, cells were counted and stained with FITC anti-CD45, PE anti-CD14, PerCP anti-CD20 and APC anti-CD3 monoclonal antibodies. Whole blood samples were assessed using standard procedure and stained accordingly for comparison with saliva. Because of low expected frequencies of lymphocytes, flow cytometric acquisition and analysis was performed by rare events detection principles. Based on stains we used and FITC threshold gating, we could discriminate major leukocyte populations (granulocytes, monocytes and lymphocytes), as well as T and B cells and measure their percentages. As the presented technique was never performed on saliva, we investigated the reliability of the method by calculating between subject and day to day within subject variation. Measurement error was assessed by calculating intra-class coefficient of correlation (ICC) for every measured parameter. We also compared saliva values to peripheral blood values. Although the substantial inter- and intraindividual variability was found in measured parameters, this is the first flow cytometric determination of salivary cell differential count so far. Predominant leukocyte population in saliva are granulocytes followed by very low frequencies of monocytes and lymphocytes. As percentages of B and T cells do not differ significantly from those of peripheral blood, saliva values could reflect changes in humoral and cellular immunity in systemic disease. It could also help us better understand characteristics of local immunoreaction, as mouth is constantly exposed to the environment and is the first barrier for microbes entering our body.

saliva; flow cytometry; leukocytes; salivary cell differential count

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