Elsevier

Colloids and Surfaces B: Biointerfaces

Volume 172, 1 December 2018, Pages 197-206
Colloids and Surfaces B: Biointerfaces

An immune magnetic nano-assembly for specifically amplifying intercellular quorum sensing signals

https://doi.org/10.1016/j.colsurfb.2018.08.033Get rights and content

Highlights

  • Small immune magnetic particles with diameter of ∼12 nm were fabricated.

  • The specific surface area of immune magnetic particles was 96.5 m2/g.

  • The distribution of antibody on the surface of magnetic particles was 25.8 μg/m2.

  • More than 3 × 108 of E. coli cells can be collected by 1 mg magnetic particles.

  • The QS signal was amplified ∼3 times than nature QS signal at total cell OD 0.08.

Abstract

Quorum sensing (QS) enables intercellular communication after bacterial cells sense the autoinducers have reached or exceeded a critical concentration. Selectively amplifying specific bacterial “quorum” activity at a lower cell density is still a challenge. Here, we propose a novel platform of immune magnetic nano-assembly to amplify specific bacterial QS signaling via improving the bioavailability of autoinducers-2 (AI-2, furanosyl borate) from sender (wide-type, WT cells) to receiver (reporter cells). Antibody coated magnetic nanoparticle (MNPAB) was fabricated with an average diameter of 12 nm and a specific surface area of 96.5 m2/g. The distribution efficiency of the antibody on the surface was 25.8 μg/m2 of magnetic nanoparticles. It was found that more than 3 × 108 of K12 serotype Escherichia coli (E. coli) reporter or WT cells were collected using 1 mg fabricated MNPAB at a saturated condition. The MNPAB not only captured E. coli WT cells but also brought them into proximity of E. coli (CT104, pCT6+pET-DsRed) reporter cells via magnetic attraction. The amplified QS signaling of the reporter cells by this immune magnetic nano-assembly was approximately 3 times higher than the nature QS signaling in cell suspension at optical density (OD) 0.08. This study foresees potential applications in amplifying specific biological QS signals based on a preprogrammed design.

Introduction

Improving bioavailability of bacterial signaling molecules is important to facilitate cell gene regulation [1] and bioactive secreta generation [2]. Extracellular signaling molecules, i.e. autoinducers, are important for bacterial communication as the cell density has reached or exceeded a critical cell density [3,4]. Furanosyl borate (autoinducer-2, AI-2) [5], a byproduct of bacterial endogenous 1-carbon metabolism [6] is such a kind of autoinducer. Those molecules have been applied externally in controlling multicellularity [7]. The quorum sensing (QS) refers to the process of generating, secreting, sensing, and transducing signaling molecules by bacteria [8]. Adding extracellular AI-2 has been applied to increase QS signaling response resulting in programmed QS activity [9]. Electronic signals were also used to mediate in vitro enzymatic synthesis of AI-2 and to further improve bacterial QS response [10]. Results from our preliminary experiments demonstrated that concentrating and focusing wide-type (WT) AI-2 releasing cells and reporter cells in a small space from a lower than typical cell density by nonspecific electrostatic absorption of magnetic nanoparticles (MNPs) improved bioavailability of bacterial AI-2, which resulted in a significant amplification of bioengineered Escherichia coli (E. coli) QS activity.

Recently, many successful methodologies have been applied to functionalize MNPs for improving cell collecting. MNPs can be coated with antibody [11,12], ovalbumin [13], vancomycin [14,15], low-molecule carbohydrates [16,17], and positively charged polymers [18,19]. Among them, Tanha et al. [12] introduced highly target-specific antibody coated MNPs to capture Staphylococcus aureus (S. aureus) cells in a mixed cell population with ∼100% efficiency and specificity. They demonstrated that antibody coated magnetic particles strongly and selectively interacted with protein A expressed on the surface of S. aureus cells, resulting in its sensitive magnetic confinement even in competition with other bacterial species [12]. Therefore, it can be speculated that using antibody coated magnetic particles for specifically collecting and concentrating WT QS sender cells and reporter cells would be beneficial to improve bioavailability of AI-2 by reporter cells.

In a previous work [20], the streptavidin-surface coated MNPs were demonstrated to effectively capture bioengineered E. coli cells with surface-expressed biotin at extra magnetic field. The resulting nano-guided cell networks revealed amplified reporter signals in an unbiased manner. Additionally, it has been found that bioengineered functional E. coli cells, i.e. E. coli (CT104 (W3110 lsrFG luxS pCT6+pET-DsRed) are extremely sensitive to AI-2, which expressed red fluorescence marker protein, Discosoma sp. red (DsRed), as a surrogate AI-2-mediated phenotypic response [9].

In this work, we reported a novel application of immune magnetic nanoparticles on amplifying QS signaling. We have designed a cell immune collector to specifically concentrate K12 serotype E. coli WT and reporter cells in a small space, which is expected to improve AI-2 bioavailability and to amplify QS signaling. Specifically, a novel immune magnetic nano-assembly was developed, and its physicochemical properties, including chemical bond composition, morphology characteristic, specific surface area, antibody surface binding efficiency, and distribution characteristic, together with its magnetism behavior were characterized. Consequently, the immune capture efficiency of WT and reporter cells by the magnetic nano-assembly was investigated. The capture properties, i.e., the specific and non-specific binding between bacterial cells and immune magnetic nano-assembly were revealed.

Section snippets

Chemicals

Ferrous sulfate heptahydrate (FeSO4·7H2O) and ferric chloride hexahydrate (FeCl3·6H2O) were purchased from the Mallinckrodt Baker, Lnc. (Phillipsburg, NJ, USA). Anti-E. coli antibody (fluorescein isothiocyanate, FITC) was purchased from the Abcam (Cambridge, MA, USA). Anti-E. coli control antibody with FITC was purchased from the Jackson ImmunoResearch Laboratories, Inc. (West Grove, PA, USA). Disodium hydrogen phosphate and sodium dihydrogen phosphate were purchased from the J. T. Baker Inc.

Physicochemical properties of MNPCOOH and MNPAB

To reveal the physicochemical properties of MNPCOOH and MNPAB, the chemical bonds, particle size, surface charge, magnetic characteristics, and binding efficiency of antibody on magnetic particles were studied systematically. The MNPCOOH was first synthesized by decomposition and recombination of iron salt crystal to generate the ferroferric oxide (Fe3O4) core which was subsequently coated by succinic acid to form stable Fesingle bondOsingle bondC bonds and COO layer in outskirt. Then, the MNPCOOH was activated by

Conclusions

We have successfully synthesized an immune magnetic nano-assembly that was able to specifically amply QS signals. The nano-assembly revealed an analogous superparamagnetic property with zeta-potential of −28.2 mV in neutral, which demonstrated that the immune magnetic nano-assembly had a good dispersibility. The average particle diameter of the immune magnetic nano-assembly was 12 nm with specific surface area of 96.5 m2/g and K12 serotype anti-E. coli antibody distribution efficiency of

Funding source

The authors gratefully acknowledge financial support from the USDA-National Institute of Food Agriculture (USDA-NIFA, #2014-67021-21585), the Defense Threat Reduction Agency (DTRA, #HDTRA1-13-0037), and the National Science Foundation (DMREF #1435957).

Acknowledgements

The authors gratefully thank Adam D. Brown in the Fischell Department of Bioengineering, Dr. Zi Teng, Dr. Sam Arnold Van Haute, and Yingying Song in the Department of Nutrition and Food Science at the University of Maryland College Park, Drs. Sz-Chian Liou and Wen-An Chiou in Department of Materials Science and Engineering at the University of Maryland College Park, and Dr. Tieren Gao in the Maryland NanoCenter for their very useful comments in this study and professional assistance for the

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