Water-powered self-propelled magnetic nanobot for rapid and highly efficient capture of circulating tumor cells

Nanosized robots with self-propelling and navigating capabilities have become an exciting field of research, attributable to their autonomous motion and specific biomolecular interaction ability for bio-analysis and diagnosis. Here, we report magnesium (Mg)-Fe3O4-based Magneto-Fluorescent Nanorobot (“MFN”) that can self-propel in blood without any other additives and can selectively and rapidly isolate cancer cells. The nanobots viz; Mg-Fe3O4-GSH-G4-Cy5-Tf and Mg-Fe3O4-GSH-G4-Cy5-Ab have been designed and synthesized by simple surface modifications and conjugation chemistry to assemble multiple components viz; (i) EpCAM antibody/transferrin, (ii) cyanine 5 NHS (Cy5) dye, (iii) fourth generation (G4) dendrimers for multiple conjugation and (iv) glutathione (GSH) by chemical conjugation onto one side of Mg nanoparticle. The nanobots propelled efficiently not only in simulated biological media, but also in blood samples. With continuous motion upon exposure to water and the presence of Fe3O4 shell on Mg nanoparticle for magnetic guidance, the nanobot offers major improvements in sensitivity, efficiency and speed by greatly enhancing capture of cancer cells. The nanobots showed excellent cancer cell capture efficiency of almost 100% both in serum and whole blood, especially with MCF7 breast cancer cells.

Predicting PY motif-mediated protein-protein interactions in the Nedd4 family of ubiquitin ligases

The Nedd4 family contains several structurally related but functionally distinct HECT-type ubiquitin ligases. The members of the Nedd4 family are known to recognize substrates through their multiple WW domains, which recognize PY motifs (PPxY, LPxY) or phospho-threonine or phospho-serine residues. To better understand protein interactor recognition mechanisms across the Nedd4 family, we report the development and implementation of a python-based tool, PxYFinder, to identify PY motifs in the primary sequences of previously identified interactors of Nedd4 and related ligases. Using PxYFinder, we find that, on average, half of Nedd4 family interactions are likely PY-motif mediated. Further, we find that PPxY motifs are more prevalent than LPxY motifs and are more likely to occur in proline-rich regions and that PPxY regions are more disordered on average relative to LPxY-containing regions. Informed by consensus sequences for PY motifs across the Nedd4 interactome, we rationally designed a focused peptide library and employed a computational screen, revealing sequence- and biomolecular interaction-dependent determinants of WW-domain/PY-motif interactions. Cumulatively, our efforts provide a new bioinformatic tool and expand our understanding of sequence and structural factors that contribute to PY-motif mediated interactor recognition across the Nedd4 family.

Comparison of methods for quantitative biomolecular interaction analysis

In order to perform good kinetic experiments, not only the experimental conditions have to be optimized, but the evaluation procedure as well. The focus of this work is the in-depth comparison of different approaches and algorithms to determine kinetic rate constants for biomolecular interaction analysis (BIA). The different algorithms are applied not only to flawless simulated data, but also to real-world measurements. We compare five mathematical approaches for the evaluation of binding curves following pseudo-first-order kinetics with different noise levels. In addition, reflectometric interference spectroscopy (RIfS) measurements of two antibodies are evaluated to determine their binding kinetics. The advantages and disadvantages of the individual approach will be investigated and discussed in detail. In summary, we will raise awareness on how to evaluate and judge results from BIA by using different approaches rather than having to rely on “black box” closed (commercial) software packages.

Plasmonic mode coupling and thin film sensing in metal–insulator–metal structures

Optical sensors based on surface plasmon resonance (SPR) in the attenuated total reflection (ATR) configuration in layered media have attracted considerable attention over the past decades owing to their ability of label free sensing in biomolecular interaction analysis, and highly sensitive detection of changes in refractive index and thickness, i.e. the optical thickness, of thin film adsorbates (thin film sensing). Furthermore, SPR is highly sensitive to the refractive index of the medium adjacent to the bare metal, and it allows for bulk sensing as well. When deposited at the metal/air interface, an adsorbed layer disturbs the highly localized, i.e. bound, wave at this interface and changes the plasmon resonance to allow for sensing in angular or wavelength interrogation and intensity measurement modes. A high degree of sensitivity is required for precise and efficient sensing, especially for biomolecular interaction analysis for early stage diagnostics; and besides conventional SPR (CSPR), several other configurations have been developed in recent years targeting sensitivity, including long-range SPR (LRSPR) and waveguide-coupled SPR (WGSPR) observed in MIM structures, referred here to by MIM modes, resulting from the coupling of SPRs at I/M interfaces, and Fano-type resonances occurring from broad and sharp modes coupling in layered structures. In our previous research, we demonstrated that MIM is better than CSPR for bulk sensing, and in this paper, we show that CSPR is better than MIM for thin film sensing for thicknesses of the sensing layer (SL) larger than 10 nm. We discuss and compare the sensitivity of CSPR and MIM for thin film sensing by using both experiments and theoretical calculations based on rigorous electromagnetic (EM) theory. We discuss in detail MIM modes coupling and anti-crossing, and we show that when a thin film adsorbate, i.e. a SL), is deposited on top of the outermost-layer of an optimized MIM structure, it modifies the characteristics of the coupled modes of the structure, and it reduces the electric field, both inside the SL and at the SL/air interface, and as a result, it decreases the sensitivity of the MIM versus the CSPR sensor. Our work is of critical importance to plasmonic mode coupling using MIM configurations, as well as to optical bio- and chemical-sensing.