Label-Free Detection of Microvesicles and Proteins by the Bundling of Gliding Microtubules

Abstract

Development of miniaturized devices for the rapid and sensitive detection of analyte is crucial for various applications across healthcare, pharmaceutical, environmental, and other industries. Here, we report on the detection of unlabeled analyte by using fluorescently labeled, antibody-conjugated microtubules in a kinesin-1 gliding motility assay. The detection principle is based on the formation of fluorescent supramolecular assemblies of microtubule bundles and spools in the presence of multivalent analytes. We demonstrate the rapid, label-free detection of CD45+ microvesicles derived from leukemia cells. Moreover, we employ our platform for the label-free detection of multivalent proteins at subnanomolar concentrations, as well as for profiling the cross-reactivity between commercially available secondary antibodies. As the detection principle is based on the molecular recognition between antigen and antibody, our method can find general application where it identifies any analyte, including clinically relevant microvesicles and proteins.

Synchronous operation of biomolecular engines

“…The recent studies on the synchronization of the biomolecular motor system will enable the precise designing of the highly computed swarming of self-propelled biomolecular motor systems (Kassem et al 2017;Sato et al 2017;Matsuda et al 2019). Moreover, the observations point towards a way of greatly improving the selectivity of many nanotechnological devices and robotic systems, such as active biosensors, sequential signaling, adaptive actuators, or analyte concentrators by optimization of these systems (Kumar et al 2012;Jia et al 2015;Chaudhuri et al 2017). However, the lifetime of the system is limited by degradation of MTs from the kinesincoated surface due to mechanical activity, reduced density of kinesin, or photodegradation (Dumont et al 2015;Keya et al 2017).…”

Curvature-Sensitive Kinesin Binding Can Explain Microtubule Ring Formation and Reveals Chaotic Dynamics in a Mathematical Model

“…Studying the mechanisms that underlie MT curling has important applications. For example, systems based on MT-kinesin gliding assays have potential uses as lab-on-a-chip medical devices, utilizing the ability to bind only selected proteins to MTs through the choice of specific cargo adapters, leading to advective transport rather than mere diffusion (Bachand et al, 2014;Chaudhuri et al, 2017). These nano-devices need to be robust for potential clinical uses, but the presence of MT rings may disrupt their design.…”

Revealing and Attenuating the Electrostatic Properties of Tubulin and Its Polymers

“…[ 1–4 ] Microtubule mechanical properties (Young's modulus ≈1 GPa; persistence length 1–10 mm [ 5–8 ] ) and ability to generate forces of up to 5 pN due to polymerization [ 9 ] are key features that enable such roles, and have led to utilization within engineered nano‐ and microelectromechanical systems (NEMS/MEMS). In tandem with molecular motors, microtubules have been employed within high efficiency rectifiers, [ 10,11 ] biosensors, [ 12–14 ] direction‐specific sorters and transporters, [ 15–18 ] force‐meters, [ 19 ] as nanopatterning agents, [ 20–22 ] and even for parallel nanocomputing. [ 23 ] Interestingly, in addition to such mechanical roles, microtubule‐based systems can also exploit the highly negative charge (47 e − ) that tubulin dimers exhibit at physiological pH values.…”