Magnetic capture from blood rescues molecular motor function in diagnostic nanodevices

Saroj Kumar,Lasse Ten Siethoff,Nuria Albet-Torres,Alf Månsson,Malin Persson

doi:10.1186/1477-3155-11-14

Abstract

BackgroundIntroduction of effective point-of-care devices for use in medical diagnostics is part of strategies to combat accelerating health-care costs. Molecular motor driven nanodevices have unique potentials in this regard due to unprecedented level of miniaturization and independence of external pumps. However motor function has been found to be inhibited by body fluids.ResultsWe report here that a unique procedure, combining separation steps that rely on antibody-antigen interactions, magnetic forces applied to magnetic nanoparticles (MPs) and the specificity of the actomyosin bond, can circumvent the deleterious effects of body fluids (e.g. blood serum). The procedure encompasses the following steps: (i) capture of analyte molecules from serum by MP-antibody conjugates, (ii) pelleting of MP-antibody-analyte complexes, using a magnetic field, followed by exchange of serum for optimized biological buffer, (iii) mixing of MP-antibody-analyte complexes with actin filaments conjugated with same polyclonal antibodies as the magnetic nanoparticles. This causes complex formation: MP-antibody-analyte-antibody-actin, and magnetic separation is used to enrich the complexes. Finally (iv) the complexes are introduced into a nanodevice for specific binding via actin filaments to surface adsorbed molecular motors (heavy meromyosin). The number of actin filaments bound to the motors in the latter step was significantly increased above the control value if protein analyte (50–60 nM) was present in serum (in step i) suggesting appreciable formation and enrichment of the MP-antibody-analyte-antibody-actin complexes. Furthermore, addition of ATP demonstrated maintained heavy meromyosin driven propulsion of actin filaments showing that the serum induced inhibition was alleviated. Detailed analysis of the procedure i-iv, using fluorescence microscopy and spectroscopy identified main targets for future optimization.ConclusionThe results demonstrate a promising approach for capturing analytes from serum for subsequent motor driven separation/detection. Indeed, the observed increase in actin filament number, in itself, signals the presence of analyte at clinically relevant nM concentration without the need for further motor driven concentration. Our analysis suggests that exchange of polyclonal for monoclonal antibodies would be a critical improvement, opening for a first clinically useful molecular motor driven lab-on-a-chip device.

Highlights

Introduction of effective point-of-care devices for use in medical diagnostics is part of strategies to combat accelerating health-care costs
In addition to serum proteins such as gelsolin, directly depolymerising the actin filaments [27], there may be binding of other blood serum components to heavy meromyosin (HMM) or actin [22]
This process is unique in that it encompasses separation steps relying on both antibody-antigen interactions, magnetic forces and the specificity of the actomyosin bond

Summary

Introduction

Introduction of effective point-of-care devices for use in medical diagnostics is part of strategies to combat accelerating health-care costs. Pumps that drive microfluidics flow demand increasingly more power the greater the miniaturization [5] and the manufacturing of the chip components becomes increasingly challenging and expensive. To overcome these problems it has been proposed that biological molecular motors, with their inherent extensive miniaturization, biodegradability and self-propelling features, may be used to transport analytes e.g. from recognition to detection chambers, achieving separation, concentration as well as certain forms of detection [6,7,8]. Several important steps towards a functional molecular motor driven diagnostic device have been realized (reviewed in [8,9,10,11,12,13]) such as: (i) attachment of antibodies to cytoskeletal microtubule [14] and actin filament [15] shuttles, followed by molecular motor-driven transportation of analytes (viruses, protein antigens etc.) bound to the antibodies, (ii) nano/ microfabrication of devices for guided transportation of the motor propelled shuttles to concentrate analytes at a detector site [6,7,16,17,18] and (iii) long-term storage of ready-to-use devices without loss of activity [19,20,21]

Methods

Results

Conclusion