Multiplexed microarrays based on optically encoded microbeads

Abstract

In recent years, there has been growing interest in optically-encoded or tagged functionalized microbeads as a solid support platform to capture proteins or nucleotides which may serve as biomarkers of various diseases. Multiplexing technologies (suspension array or planar array) based on optically encoded microspheres have made possible the observation of relatively minor changes in biomarkers related to specific diseases. The ability to identify these changes at an early stage may allow the diagnosis of serious diseases (e.g. cancer) at a time-point when curative treatment may still be possible. As the overall accuracy of current diagnostic methods for some diseases is often disappointing, multiplexed assays based on optically encoded microbeads could play an important role to detect biomarkers of diseases in a non-invasive and accurate manner. However, detection systems based on functionalized encoded microbeads are still an emerging technology, and more research needs to be done in the future. This review paper is a preliminary attempt to summarize the state-of-the-art concerning diagnostic microbeads; including microsphere composition, synthesis, encoding technology, detection systems, and applications.

Spectrally encoded microspheres for immunofluorescence analysis

“…The multiplex measurements can be based on using a number of various dyes and several different fluorescence intensities of one dye. 47 For example, in xMAP Technology (Luminex) microspheres are internally dyed with two different fluorophores in different concentrations, producing a 100-member array of spectrally distinct microsphere sets. 48 A large number of fluorophores with different emission peaks opens up wide opportunities for multiplex detection.…”

Lifetime encoding in flow cytometry for bead-based sensing of biomolecular interaction

“…Flow cytometry (FCM) is a high-throughput screening (HTS) method for the optical analysis of a large number of either single cells or microbead populations in a flow 1,2 . This multiparametric fluorescence technique commonly relies on fluorophores for optical encoding and analyte quantification through spectral and intensity information [3][4][5][6][7] and is well-established in many analytical and research applications in medical diagnostics, biology, and food analysis [8][9][10][11] . FCM is particularly attractive for the readout of multiplexed bioassays and the HTS of biomarkers e.g.…”

“…Typical encoding schemes utilize spectral and/or intensity codes. Spectral codes can, however, suffer from spectral overlap of the emission bands of encoding luminophores, limiting the number of distinguishable codes and often requiring signal compensation and cross talk corrections 10,19 . Intensity encoding is subject to leaking and photobleaching of the encoding fluorophores and is affected by fluctuations in the excitation light intensity which can lead to systematic errors in code assignment.…”

“…To increase the number of distinguishable codes in FCM and circumvent these possible drawbacks of spectral multiplexing in the intensity domain, the characteristic fluorescence lifetime (LT) of a fluorophore can be exploited as an additional detection or encoding parameter. An additional advantage of LT measurements is the independence of LT on fluorophore concentration and excitation light intensity which, together with decreasing costs of pulsed light sources and fast detectors, renders time-resolved fluorescence detection increasingly attractive for imaging and sensing applications 4,10,[20][21][22] . LT-FCM has already been considered before as well 5 , and was mostly addressed by combining FCM with fast frequency domain LT techniques compatible with the high-throughput conditions in FCM 5,6,[22][23][24][25][26][27] .…”

Trends of Bead Counting-Based Technologies Toward the Detection of Disease-Related Biomarkers

“…It should be noted that in such strategies, the concentration of target can only be obtained by substituting the experimental data into the standard curve established by a gradient of standard samples with known concentrations. Basically, there are two representative types of statistical bead counting strategies: the solid-state planar arrays and the liquid-state suspension arrays, both of which have been applied widely in the detection of disease-related biomolecules (Parsa et al, 2018 ; Vafajoo et al, 2018 ). In the solid-state planar arrays, the beads are attached to a solid substrate after target capturing and fluorescence signal accumulation, and the fluorescence signals of the beads are monitored after the washing procedure (Sukhanova and Nabiev, 2008 ).…”