Emerging Technologies to Increase Ligand Binding Assay Sensitivity

Abstract

Abstract. Ligand binding assays (LBAs) have been the method of choice for protein analyte measurements for more than four decades. Over the years, LBA methods have improved in sensitivity and achieved larger dynamic ranges by using alternative detection systems and new technologies. As a consequence, the landscape and application of immunoassay platforms has changed dramatically.

Highly sensitive ligand-binding assays in pre-clinical and clinical applications: immuno-PCR and other emerging techniques

“…While the relatively young technology of DNA-enhanced LBAs has previously been surveyed, [44][45][46][47][48][49][50][51][52] we will here discuss this technology in the context of other emerging techniques for high-sensitivity analyses of biomacromolecules with an emphasis on case studies from preclinical and clinical sample testing in support of pharmaceutical drug development programs. 57 54 Specifically, we discuss technology platforms which have found their way into routine ( pre)clinical sample testing, DNA-enhanced immunoassays, electrochemiluminescence (ECL), 55 single molecule counting (SMC) 56 and a brief selection of other emerging LBA technologies.…”

“…An overview of specific experimental details of these technologies is given in Table 1. 57 Similar, IPCR is gaining an increased significance for pharmaceutical development processes (chapter 3.3), while the related nucleic acidenhanced proximity ligation currently appears to have an emphasis on biomarker discovery (chapter 3.4). This technique has already been reviewed extensively 55,96-100 and we will therefore focus here on selected recent case studies.…”

“…[137][138][139] These issues also apply for alternate approaches, such as bead-based IPCR assays 49,91,140,141 or chemically activated glass surfaces designed for automated processing. 57,150,154 This not only enables pre-clinical toxicology studies to support investigational new drug (IND) programs but also to realize fully validated IPCR protocols for sample testing support under GLP regime. 126 For instance, highly sensitive quantitative Immuno-PCR (qIPCR) 66 assays, commercially available since the early 2000 years, contain pre-synthesized antibody-DNA conjugates.…”

Platforms and Techniques Used for Biomarker Assays: Where are We Now?

“…However, as the need to develop ever more sensitive assays increases, problems with developing selective assays becomes more challenging. Historically, photometric LBAs were the most commonly used platform for both commercial and bespoke assays, however, In addition, various vendors have been developing proprietary ultrasensitive instrumentation to allow quantitation of very low abundance biomarkers, including: MSD platform (S-Plex), Singulex and Quanterix Simoa with the various platforms providing different sensitivities depending on the analyte of interest [2,3]. Although these platforms can provide high sensitivity assays, they also suffer from additional challenges associated with selectivity from individual samples due to the low levels of analyte that are being detected and often lead to a trade-off between assay sensitivity and overall selectivity.…”

Detection and Digital Resolution Counting of Nanoparticles with Optical Resonators and Applications in Biosensing

“…Recently, ultrasensitive assay technologies with "digital" analyte precision have been introduced, and summarized in reviews [39,40]. For example, the Simoa™ system by Quanterix [41], uses an enzymatically amplified fluorescent reporter attached to antibody-functionalized micron-scale magnetic beads isolated in 50-fl reaction chambers to achieve non-multiplexed fM-scale detection limits with a more complex protocol than enzyme-linked immunosorbant ELISA.…”