Ravara Barbara, Zampieri Sandra, Kern Helmut, Carraro Ugo
CIR-Myo, Interdepartmental Research Center of Myology, University of Padova, Italy.
Department of Biomedical Sciences (DSB), University of Padova, Italy.
Eur J Transl Myol. 2019 Dec 10;29(4):8713. doi: 10.4081/ejtm.2019.8713. eCollection 2019 Oct 29.
Use of saliva in clinical studies are increasing to identify methods less invasive than blood sampling in search for systemic changes of biomarkers related to physical activity, aging, late aging and rehabilitation. The consensus is that the diagnostic value of whole saliva is compromised by the presence of blood, but we are looking at the contamination as a major opportunity for non-invasive analyses of serological biomarkers. The aim of this preliminary study was to evaluate the presence of serum in mouth fluids of healthy seniors and the eventual changes after a modest trauma, i.e., tooth brushing. Seven heathy persons, aged more than 65 years, drooling saliva in a test tube provided the fluids for the analyses. After low speed centrifugation, small aliquots of supernatants were frozen in liquid nitrogen and stored at -80° until use. Aliquots were thawed and used for quantification by the Lowry method of total proteins and by colorimetric ELISA of serum albumin, fibrinogen and lysozyme. Hemoglobin content was quantified by Spectrophotometry. Adjustment of saliva dilution, after a preliminary test, increased the homogeneity of the analytes' content determined by colorimetric ELISA. The control reference to judge the quantity of serum in saliva was a pool of sera from age-matched healthy persons. Saliva collected from the seven healthy elderly person before and after tooth-and-gum, brushing presented measurable amount of the analytes, including fibrinogen, a minor component of the pooled sera. Tooth brushing did not induced statistically significant difference in analytes' contents, suggesting that a measurable blood contamination is a frequent event in elderly persons. In conclusion, fibrinogen analysis in saliva is a promising approach to quantify serological biomarkers by a non-invasive procedure that will increase acceptability and frequency of analyses during follow-up in aging and rehabilitation.
在临床研究中，唾液的使用正在增加，以寻找比采血侵入性更小的方法，来探寻与身体活动、衰老、老年晚期和康复相关的生物标志物的全身变化。目前的共识是，全唾液的诊断价值会因血液的存在而受到影响，但我们将这种污染视为对血清生物标志物进行非侵入性分析的一个主要契机。这项初步研究的目的是评估健康老年人唾液中血清的存在情况，以及在轻微创伤（即刷牙）后的最终变化。七名年龄超过65岁的健康人，将唾液滴入提供的试管中，以供分析使用。低速离心后，将少量上清液在液氮中冷冻，并在-80°下保存直至使用。取出部分解冻后，用于通过Lowry法对总蛋白进行定量，并通过比色ELISA法对血清白蛋白、纤维蛋白原和溶菌酶进行定量。通过分光光度法对血红蛋白含量进行定量。经过初步测试后，调整唾液稀释度，提高了比色ELISA法测定的分析物含量的均匀性。判断唾液中血清量的对照参考是来自年龄匹配的健康人的混合血清。在刷牙前后从这七名健康老年人收集的唾液中，呈现出可测量的分析物含量，包括纤维蛋白原，这是混合血清中的次要成分。刷牙并未导致分析物含量出现统计学上的显著差异，这表明可测量的血液污染在老年人中是常见现象。总之，唾液中的纤维蛋白原分析是一种很有前景的方法，可通过非侵入性程序对血清生物标志物进行定量，这将提高在衰老和康复随访期间分析的可接受性和频率。
