Rapid Biomarker Detection Research Articles

SPR detection of cardiac troponin T for acute myocardial infarction

A surface plasmon resonance (SPR) sensor developed for the rapid, sensitive and specific detection of cardiac troponin T (cTnT) in serum samples is reported in this work. An extensive optimisation of assay parameters was conducted to achieve optimal detection strategy. Both direct and sandwich immunoassay formats were investigated and optimised. The response obtained was enhanced further by the use of gold nanoparticles (AuNPs) conjugated to the anti-cTnT detection antibody. A regeneration method was developed to enable the reuse of the SPR sensor for multiple sample application. The SPR immunosensor showed good reproducibility for cTnT detection in the concentration range of 25–1000ngmL−1 and 5–400ngmL−1 for the direct and sandwich assays in buffer, respectively. The linear regression analysis was performed and R2 value was found as 0.99 for both assays. In order to optimise the sensor for serum analysis, nonspecific binding of serum proteins was reduced through the use of additives in the dilution buffer. To achieve greater sensitivity, the performance of the cTnT immunosensor sandwich assay in human serum was evaluated using non-modified and AuNP modified detector antibodies. A detection limit (LOD) for the immunosensor in 50% serum was assessed as 5ngmL−1 cTnT for the standard sandwich assay and 0.5ngmL−1 cTnT when using AuNP conjugated detector antibodies with a linear dynamic range of 0.5–40ngmL−1. The dissociation constant was found as 3.28 × 10−9M using Langmuir binding model which indicates high affinity between cTnT and its antibody. The proposed SPR immunosensor has a promising potential to be developed for point-of-care testing for the early diagnosis of acute myocardial infarction (AMI). This method can also be used for the rapid detection of biomarkers in central nervous system diseases.

Abstract 3220: A microfluidic, multi-analyte, high sensitivity, immunoassay platform for rapid biomarker detection.

Abstract Traditionally, protein analytes or biomarkers have been measured individually in ELISAs. However, it is becoming increasingly clear that multiple markers are associated with multivariate diseases like cancer. Despite demand, the adoption of multi-analyte biomarker tests in clinical research has been severely limited for many reasons including technical concerns regarding assay reproducibility, time and the labor-intensive nature of assay panel development, and reported non-correlation with conventional ELISA data. To address this issue, we have developed a novel fully automated desktop immunoassay platform that enables simultaneous interrogation of four analytes in eight individual samples on a single disposable microfluidic cartridge in under an hour. We have devised a unique approach to immobilize antibodies that results in assays with high sensitivities (femtomolar) and broad dynamic ranges (>4 logs). Twenty microliters of patient sample are split into parallel channels containing individual immunoassays. By segregating each assay to a discrete location, the multi-analyte assay mimics single-analyte ELISAs, thus enabling each assay to operate under its optimal conditions. This methodology combines the specificity of a traditional ELISA immunoassay, where the sample is assayed by a singular pair of capture and detect antibodies eliminating cross-reactivity and other negative interactions from other antibody pairs while offering the benefits of a multiplexed antigen analysis. We describe the system's design, speed, sensitivity and reproducibility using data generated with a custom-built 4-analyte pro-inflammatory panel cartridge (IL-1β, IL-6, IL-10 and TNFα) on clinical samples. We demonstrate that our assay system is ideal for quantitative analysis of soluble protein markers in clinical research and in clinical trials. Citation Format: Rajiv Pande, John H. Leamon, Marty Putnam, M Pitta, D Svancara, J Tu. A microfluidic, multi-analyte, high sensitivity, immunoassay platform for rapid biomarker detection. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3220. doi:10.1158/1538-7445.AM2013-3220

Detection of Orexin A Neuropeptide in Biological Fluids Using a Zinc Oxide Field Effect Transistor

Biomarkers which are indicative of acute physiological and emotional states are studied in a number of different areas in cognitive neuroscience. Currently, many cognitive studies are conducted based on programmed tasks followed by timed biofluid sampling, central laboratory processing, and followed by data analysis. In this work, we present a sensor platform capable of rapid biomarker detection specific for detecting neuropeptide orexin A, found in blood and saliva and known as an indicator of fatigue and cognitive performance. A peptide recognition element that selectively binds to orexin A was designed, characterized, and functionalized onto a zinc oxide field effect transistor to enable rapid detection. The detection limit using the sensor platform was sub-picomolar in water, and picomolar to nanomolar levels in saliva and serum. The transistor and recognition element sensor platform can be easily expanded, allowing for multiple biomarkers to be detected simultaneously, lending itself to complex biomarker analysis applicable to rapid feedback for neuroscience research and physiological monitoring.

Reagent-Loaded Cartridges for Valveless and Automated Fluid Delivery in Microfluidic Devices

An important problem in the life sciences and in health care is simple and rapid detection of biomarkers. Although microfluidic devices are potentially useful in addressing this problem, current techniques for automating fluid delivery--which include valves and electroosmosis--require sophisticated microfabrication of the chip, bulky instrumentation, or both. In this paper, we describe a simple and reliable technique for storing and delivering a sequence of reagents to a microfluidic device. The technique is low-cost, requires minimal user intervention, and can be performed in resource-poor settings (e.g., outside of a laboratory) in the absence of electricity and computer-controlled equipment. In this method, cartridges made of commercially available tubing are filled by sequentially injecting plugs of reagents separated by air spacers. The air spacers prevent the reagents from mixing with each other during cartridge preparation, storage, and usage. As an example, we used this "plug-in cartridge" technology to complete a solid-phase immunoassay in a microchannel in 2 min with low-nanomolar sensitivity and demonstrate the diagnosis of HIV in 13 min.

Field-portable, automated pyrolysis-GC/IMS system for rapid biomarker detection in aerosols: A feasibility study

A prototype automated pyrolysis-gas chromatography/ion mobility spectrometry (Py-GC/IMS) instrument was developed for (bio)aerosol characterization. The system combines a commercially available, hand-held GC/IMS device with a specially built platinum wire grid heater, a 1-in.-diam. quartz microfiber filter and a 60-l/min air pump. The prototype Py-GC/IMS system can be operated in stand-alone mode or in series with a particle concentrator. Fully automated collection/desorption/pyrolysis of aerosols and other particulate matter can be performed at repetition rates of up to 60 h−1 by means of a special remote control and display software package. The feasibility of detecting submicrogram quantities of Bacillus endospores with the use of picolinic acid and pyridine as biochemical marker compounds for the characteristic dipicolinic acid moiety in spore cell walls was demonstrated by laboratory experiments as well as preliminary field tests. Other particulate matter that could be collected and analyzed includes allergens such as pollen or home dust, as well as a broad range of bioaerosols and reaerosolized organics like explosives or drugs. In addition to its potential use as a screening device for the presence of specific classes of aerosol components, the Py-GC/IMS system has been demonstrated to retain its capability to detect and identify a broad range of volatile and semivolatile organic compounds. © 1997 John Wiley & Sons, Inc. Field Analyt Chem Technol 1: 295–305, 1997