Using silicon-based biosensors to detect host protein signatures capable of differentiating between bacterial and viral infection at point-of-care

世界卫生组织预测，到2050年，抗菌素耐药性(AMR)的上升将夺走多达1000万人的生命[1]。导致AMR出现的关键因素之一是缺乏能够区分细菌感染和病毒感染的快速诊断设备。细菌培养试验是目前的“黄金标准”，然而它存在着各种缺点，如阳性结果的时间较长和样本污染的风险[2]。这导致临床医生要么在等待确认时推迟抗生素治疗，要么在严重感染的情况下过度开广谱抗生素。因此，了解到大约50%的抗生素被不当开出也就不足为奇了。理想的诊断设备应该是快速、高灵敏度和高特异度的现场可用诊断设备，然而，当前诊断技术的挑战使这些特征很难实现。首先，它们主要依赖于基于光学的检测方法，这种方法通常需要庞大而昂贵的实验室设备，因此经常使这些分析在护理地点无法获得[4]。此外，直接检测致病病原体可能是不可能的，因为一些病原体具有令人难以置信的侵袭性或无法接触到，特别是在结核病的情况下。最后，确认细菌感染并不排除引起病毒感染，反之亦然，这在上呼吸道感染中是一种非常常见的动态[5]。宿主蛋白签名可以绕过所有这些挑战，各种宿主蛋白签名已经在文献中被引用。奥维德等人。(2015)发现了一个由C-反应蛋白(CRP)、肿瘤坏死因子相关的凋亡诱导配体(TRAIL)和干扰素-γ诱导蛋白-10(IP-10)组成的3宿主蛋白特征，能够准确区分细菌感染和病毒感染[6]。然而，据估计，超过90%的已发现宿主生物标记物尚未在护理点成功应用[7]。将宿主蛋白质签名与已经利用的互补金属氧化物半导体技术(CMOS)(用于制造日常电子产品的领先技术)相结合，将使这种设备变得难以置信地廉价和可扩展[8]。CMOS技术还与离子敏感场效应晶体管(ISFET)兼容，它可以用来检测由于抗原-抗体结合引起的表面电荷的变化，从而实时检测和定量宿主蛋白。研究问题：-什么特定的宿主蛋白特征能够以高灵敏度和特异性区分细菌和病毒感染？-芯片上固定的免疫分析的灵敏度和特异度与“金标准”蛋白质检测技术相比如何？-需要什么技术来将主要抗体固定在ISFET传感器阵列上？-该特定蛋白特征的芯片检测极限是多少？目的：-本项目旨在开发一种免疫分析能够区分细菌感染和病毒感染的宿主蛋白特征。-然后，特定的免疫分析将被转换到硅基生物传感器上，并将建立芯片上的检测极限。-最后，该化验将与临床样本进行验证，并用于提供有关患者感染状况的量化数据。参考文献[1]M.E.A.de Kraker、A.J.Stewardson和S.Harbarth，《到2050年每年是否有1,000万人因抗菌素耐药性而死亡？》，《公共科学图书馆医学》，第13卷，第11期，E1002184页，2016年11月，DOI：10.1371/JOURNAL.PMED.1002184。[2]J.C.Craig等人，《诊断发烧儿童严重细菌感染的临床症状和体征的准确性：15781例发热疾病的前瞻性队列研究》，《英国医学杂志》(临床研究版)，第340卷，第340页。第7754号，第1015页，2010年5月，DOI：10.1136/BMJ.C1594.[3]C.朱利亚诺，C.R.帕特尔，和

The World Health Organisation has predicted that the rise of antimicrobial resistance (AMR) will claim up to 10 million lives by the year 2050[1]. One of the key factors driving AMR emergence is the lack of a rapid diagnostic device capable of differentiating between bacterial and viral infection. A bacterial culture test is the current "gold standard", however it presents with various draw backs such as long time-to-positive results and risk of sample contamination[2]. This leads to clinicians either delaying antibiotic treatment as they wait for confirmation or overprescribing broad spectrum antibiotics in cases of severe infection. It is then unsurprising to learn that around 50% of antibiotics are inappropriately prescribed[3]. The ideal diagnostic device would be rapid and available at point-of-care with high sensitivity and specificity, however challenges with current diagnostics make these characteristics difficult to achieve. Primarily, they mostly rely on optical-based methods of detection which usually require bulky and expensive laboratory equipment and so often makes these assays unavailable at point-of-care[4]. Additionally, direct detection of the causative pathogen may not be possible as some pathogens are incredibly invasive or inaccessible, especially in cases of tuberculosis. Finally, confirmation of a bacterial infection does not rule out a causative viral infection and vice versa which is a very common dynamic in upper respiratory infections[5].Host protein signatures could bypass all these challenges and various host protein signatures have already been cited in literature. Oved et al. (2015) discovered a 3 host protein signature composed of C-reactive protein (CRP), TNF-related apoptosis-inducing ligand (TRAIL) and Interferon gamma-induced protein-10 (IP-10) which is able to accurately differentiate between bacterial and viral infection[6]. However, it is estimated that over 90% of discovered host biomarkers have not yet been applied successfully at point-of-care[7]. Combining host protein signatures with already leveraged complementary metal-oxide semiconductor technology (CMOS) (the leading technology used to manufacture everyday electronics) would make such devices incredibly cheap and scalable[8]. CMOS technology is also compatible with ion sensitive field effect transistors (ISFETs), which can be used to detect changes in surface charge due to antigen-antibody binding and hence detect and quantify host proteins in real time.Research questions:-What specific host protein signature is able to differentiate between bacterial and viral infection with high sensitivity and specificity? -How does the sensitivity and specificity of the immunoassay immobilised on-chip compare to the "gold standard" protein detection technique?-What techniques are required to immobilise primary antibodies onto an array of ISFET sensors? -What is the on-chip limit of detection for that specific protein signature?Aims:-This project aims to develop an immunoassay specific to a host protein signature capable of differentiation between bacterial and viral infection. -The specific immunoassay will then be translated onto a silicon-based biosensor and the on-chip limit of detection will be established. -Finally, the assay will be validated against clinical samples and used to provide quantitative data around the infection state of the patient. References [1] M. E. A. de Kraker, A. J. Stewardson, and S. Harbarth, "Will 10 Million People Die a Year due to Antimicrobial Resistance by 2050?," PLOS Medicine, vol. 13, no. 11, p. e1002184, Nov. 2016, doi: 10.1371/JOURNAL.PMED.1002184.[2] J. C. Craig et al., "The accuracy of clinical symptoms and signs for the diagnosis of serious bacterial infection in young febrile children: prospective cohort study of 15 781 febrile illnesses," BMJ (Clinical research ed.), vol. 340, no. 7754, p. 1015, May 2010, doi: 10.1136/BMJ.C1594.[3] C. Giuliano, C. R. Patel, and