Oak Ridge Institute for Science and Education, Oak Ridge, TN, USA.
J Immunol Methods. 2011 Feb 1;364(1-2):83-93. doi: 10.1016/j.jim.2010.11.005. Epub 2010 Nov 18.
Saliva has an important advantage over serum as a medium for antibody detection due to non-invasive sampling, which is critical for community-based epidemiological surveys. The development of a Luminex multiplex immunoassay for measurement of salivary IgG and IgA responses to potentially waterborne pathogens, Helicobacter pylori, Toxoplasma gondii, Cryptosporidium, and four noroviruses, involved selection of antigens and optimization of antigen coupling to Luminex microspheres. Coupling confirmation was conducted using antigen specific antibody or control sera at serial dilutions. Dose-response curves corresponding to different coupling conditions were compared using statistical tests. Control proteins in the specific antibody assay and a separate duplex assay for total immunoglobulins G and A were employed to assess antibody cross-reactivity and variability in saliva composition. 200 saliva samples prospectively collected from 20 adult volunteers and 10 paired sera from a subset of these volunteers were used to test this method. For chronic infections, H. pylori and T. gondii, individuals who tested IgG seropositive using commercial diagnostic ELISA also had the strongest salivary antibody responses in salivary antibody tests. A steep increase in anti-norovirus salivary antibody response (immunoconversion) was observed after an episode of acute diarrhea and vomiting in a volunteer. The Luminex assay also detected seroconversions to Cryptosporidium using control sera from infected children. Ongoing efforts involve further verification of salivary antibody tests and their application in larger pilot community studies.
唾液相对于血清作为抗体检测的介质具有重要优势，因为唾液是一种非侵入性的采样方式，这对于基于社区的流行病学调查至关重要。为了测量唾液 IgG 和 IgA 对潜在水源性病原体（如幽门螺杆菌、刚地弓形虫、隐孢子虫和四种诺如病毒）的反应，我们开发了一种 Luminex 多重免疫分析方法，该方法涉及抗原的选择和抗原与 Luminex 微球的偶联优化。使用抗原特异性抗体或对照血清在系列稀释度下进行偶联确认。使用统计检验比较了对应于不同偶联条件的剂量反应曲线。在特异性抗体检测中使用了控制蛋白和单独的总 IgG 和 IgA 双检测，以评估抗体的交叉反应性和唾液成分的可变性。前瞻性地从 20 名成年志愿者中收集了 200 份唾液样本，以及从这些志愿者中的一部分中收集了 10 对配对血清，用于测试该方法。对于慢性感染，如幽门螺杆菌和刚地弓形虫，使用商业诊断 ELISA 检测 IgG 血清阳性的个体在唾液抗体检测中也具有最强的唾液抗体反应。在一名志愿者出现急性腹泻和呕吐后，抗诺如病毒唾液抗体反应（免疫转化）急剧增加。Luminex 分析还使用感染儿童的对照血清检测到了隐孢子虫的血清转化。目前正在进行进一步验证唾液抗体检测及其在更大规模的试点社区研究中的应用。
