Probing basophil function in microfluidic systems for allergic disease diagnosis

项目总结 嗜碱性细胞在过敏性疾病中扮演着重要的角色：它们的激活越来越多地被用于食物 变态反应的诊断，以及它们向组织中的迁移与变态反应的发生有关 过敏反应。然而，由于嗜碱性粒细胞很少见，它们在诊断中的研究和利用较少。 比其他白细胞，特别是在体外微流控系统中。这个项目的总体目标是 是通过开发使微流控技术能够更好地量化嗜碱性粒细胞来填补这一空白 功能，并以食物过敏为模型疾病。 食物过敏已经达到流行的程度，并已成为 医疗负担。常见的体外过敏试验缺乏足够的准确性。口腔食物挑战赛， 食物过敏评估的黄金标准，通常不执行，因为它将患者置于 有过敏反应的风险。因此，食物过敏通常只有在出现以下不良反应后才会被发现 可能会危及生命。此外，没有可靠的相关性表明 过敏性反应和最低剂量的过敏原。用来预测过敏反应风险的指标 病人基本上是不存在的。 我们的长期目标是通过开发过敏诊断测试来应对这些挑战 准确、安全、快速和可访问的指标，以及2)更好地对高危患者进行分层 过敏反应。这项试点拨款的总体目标是检验以下假设：1)阻抗 嗜碱性细胞在激活时的变化，这暴露出带负电荷的外化颗粒 蛋白多糖，是一种准确和简单的无标记的嗜碱性细胞激活指示剂，以及2) 嗜碱性细胞迁移和分泌情况与反应严重程度相关。我们的方法包括 使用临床证实的和特征良好的花生过敏受试者的全血样本 和健康控制，并利用我们跨学科团队在创新方面的专业知识 微流控技术和人体免疫学。我们将：(目标1)设计和验证无标签 食物过敏原存在下的全血阻抗法嗜碱粒细胞活化试验 体内，以及(目标2)设计和验证耦合的嗜碱性细胞迁移和多路分泌 体外试验，以确定与患者过敏反应病史的相关性。

PROJECT SUMMARY Basophils play important roles in allergic diseases: their activation is increasingly used for food allergy diagnosis, and their migration into tissues has been correlated with the onset of anaphylaxis. However, since basophils are rare, they are less studied and utilized for diagnostics than other leukocytes, especially in ex vivo microfluidic systems. The overall goal of this project is to fill this gap by developing enabling microfluidic technologies to better quantify basophil function, and by using food allergy as a model disease. Food allergy has reached epidemic proportions and has become a significant source of healthcare burden. Common in vitro allergy tests lack sufficient accuracy. Oral food challenge, the gold standard for food allergy assessment, is often not performed as it places the patient at risk of anaphylaxis. As such, food allergy is often identified only after an adverse reaction that could be life-threatening. In addition, there is no reliable correlation between the onset of anaphylaxis and a minimum offending dose of allergen. Metrics to predict anaphylaxis risk in a patient are essentially non-existent. Our long-term goal is to address these challenges by developing 1) allergy diagnostic tests that are accurate, safe, rapid, and accessible, and 2) metrics to better stratify patients at high risk of anaphylaxis. The overall objective of this pilot grant is to test the hypotheses that: 1) impedance changes in basophils upon activation, which exposes negatively-charged exteriorized granule proteoglycans, is an accurate and simple label-free indicator for basophil activation, and 2) basophil migration and secretion profile correlates with reaction severity. Our approach includes using whole blood samples from clinically-proven and well-characterized peanut allergic subjects and healthy controls, and leveraging our interdisciplinary team's expertise in innovative microfluidic technology and human immunology. We will: (Aim 1) design and validate a label-free impedance-based basophil activation test using whole blood in the presence of food allergen ex vivo, and (Aim 2) design and validate a coupled basophil migration and multiplexed secretion assay ex vivo to determine the correlation with patient history of anaphylaxis.