Research ArticleMEDICAL ROBOTS

Hybrid biomembrane–functionalized nanorobots for concurrent removal of pathogenic bacteria and toxins

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Science Robotics  30 May 2018:
Vol. 3, Issue 18, eaat0485
DOI: 10.1126/scirobotics.aat0485
  • Fig. 1 Preparation and characterization of RBC-PL-robots.

    (A) Schematic of biomimetic robots for binding and removal of threatening biological agents. (B) Preparation of RBC-PL-robots: (i) The gold surface of the nanowire (AuNW) robots was modified with MPA; (ii) hybrid membranes were prepared by fusion of RBC membranes and PL membranes (using 1:1 protein weight ratio), and the resulting hybrid membranes were used to coat the MPA-modified nanorobots; (iii) after 5-min sonication, the RBC-PL-robots were obtained. (C) SEM images of a bare AuNW robot without hybrid membrane coating (top) and an RBC-PL-robot (bottom). (D) The measured weight of protein content on bare robots and RBC-PL-robots (both 20 mg ml−1) stored in 1× PBS at 4°C for 24 hours. UD, undetectable. Error bars represent the SD from three different measurements. (E) SDS–polyacrylamide gel electrophoresis analysis of proteins present on the RBC-PL-vesicles and the RBC-PL-robots. The samples were run at equal protein content and stained with Coomassie blue. The RBC-PL-robots used in (D) and (E) were exposed to the acoustic field for 5 min before performing the protein analysis. (F) Optical (i) and fluorescent (ii to iv) images of a group of RBC-PL-robots, in which the RBC membranes were labeled with DiD dye (red) and the PL membranes were labeled with FITC (green). Overlay of the DiD and FITC channels is shown in (iv) (yellow).

  • Fig. 2 Propulsion performance and anti-biofouling property of RBC-PL-robots.

    Tracking trajectories showing the propulsion of bare robots (A) and RBC-PL-robots (B) in water (i) and after 0- and 1-hour incubation in blood (ii and iii, respectively). (C) Comparison of the speed of bare robots with RBC-PL-robots in water and after 0- and 1-hour incubation in blood. The acoustic nanorobots were propelled using a frequency of 2.66 MHz and a voltage of 2.0 V. Error bars estimated as a triple of SD (n = 3).

  • Fig. 3 Binding and isolation of PL-adhering pathogens by RBC-PL-robots.

    (A) SEM image of an MRSA USA300 bacterium attached to an RBC-PL-robot. (B) Microscopic images showing the binding of an MRSA USA300 bacterium to an RBC-PL-robot: bright-field image (top) and fluorescence image showing the DAPI-stained bacterium (bottom). (C) Normalized fluorescence intensity of DAPI-stained MRSA USA300 bacteria retained on (i) PBS (no robots), (ii) bare robots, (iii) RBC-robots (without PL membranes), (iv) RBC-PL-robots under a static condition (without US), (v) RBC-PL-vesicles, (vi) US-propelled RBC-PL-robots, and (vii) PL-robots (without RBC membranes). PBS, bare robots, RBC-robots, and RBC-PL-vesicles were used as negative controls; PL-robots were used as a positive control. Error bars were estimated as a triple of SD (n = 3). Reaction time, 5 min. Biomimetic robots, 10 mg ml−1. MRSA USA300 bacteria, 5 × 108 CFU ml−1.

  • Fig. 4 Binding and neutralization of α-toxin and other PFTs by RBC-PL-robots.

    (A) Images of centrifuged 5% RBC solution after incubation with α-toxin in (i) PBS, (ii) static RBC-PL-robots, (iii) US-propelled RBC-PL-robots, and (iv) US-propelled RBC-robots. PBS without α-toxin (v) was used as a control. (B) Hemolysis quantification of the samples shown in (A). (C) Images of centrifuged 5% RBC solution after incubation with PFTs (50 μl) produced by MRSA bacteria (after 8-hour incubation) in (i) PBS, (ii) static RBC-PL-robots, (iii) US-propelled RBC-PL-robots, and (iv) US-propelled RBC-robots. PBS without PFTs (v) was used as a control. (D) Hemolysis quantification of the samples shown in (C). Error bars estimated as a triple of SD (n = 3). Treatment time with samples, 5 min; hemolysis incubation time (5% RBC solution plus PFTs), 30 min at 37°C.

  • Fig. 5 In situ concurrent removal of MRSA bacteria and MRSA-secreting PFTs by RBC-PL-robots.

    (A) Schematic of RBC-PL-robots for bacteria targeting and PFT neutralization. (B) Optical density values (OD600) of MRSA bacteria obtained before and after treatment (blue, without robots; red, RBC-PL-robots). (C) Relative hemolysis percentages of PFTs obtained before and after treatment (blue, without robots; red, RBC-PL-robots). Error bars estimated as a triple of SD (n = 3). (D) MRSA bacterial growth curves (indicated by OD600) versus incubation time for nontreated bacteria and bacteria treated with RBC-PL-robots. (E) Corresponding relative hemolysis curves versus incubation time for nontreated bacteria and bacteria treated with RBC-PL-robots. Arrows indicate first measurement right after treatment.

Supplementary Materials

  • robotics.sciencemag.org/cgi/content/full/3/18/eaat0485/DC1

    Fig. S1. SEM images of bare AuNW robots along with the corresponding EDX analysis showing the Au nanorobots’ body.

    Fig. S2. SEM images of MRSA USA300 bacteria attached to RBC-PL-robots.

    Fig. S3. Absorbance spectra of oxyhemoglobin after incubation with commercial purified α-toxin under different conditions.

    Fig. S4. Absorbance spectra of oxyhemoglobin after incubation with a cocktail of PFTs produced by MRSA bacteria under different conditions.

    Fig. S5. Characterization of bare Au/Ni/Au NW robots.

    Fig. S6. Experimental protocol followed to perform the combined application of the RBC-PL-robots: MRSA targeting and PFT neutralization.

    Movie S1. Propulsion performance of bare robots in water and whole blood under a US field.

    Movie S2. Propulsion performance of RBC-PL-robots in water and whole blood under a US field.

  • Supplementary Materials

    Supplementary Material for:

    Hybrid biomembrane–functionalized nanorobots for concurrent removal of pathogenic bacteria and toxins

    Berta Esteban-Fernández de Ávila, Pavimol Angsantikul, Doris E. Ramírez-Herrera, Fernando Soto, Hazhir Teymourian, Diana Dehaini, Yijie Chen, Liangfang Zhang,* Joseph Wang*

    *Corresponding author. Email: josephwang{at}ucsd.edu (J.W.); zhang{at}ucsd.edu (L.Z.)

    Published 30 May 2018, Sci. Robot. 3, eaat0485 (2018)
    DOI: 10.1126/scirobotics.aat0485

    This PDF file includes:

    • Fig. S1. SEM images of bare AuNW robots along with the corresponding EDX analysis showing the Au nanorobots’ body.
    • Fig. S2. SEM images of MRSA USA300 bacteria attached to RBC-PL-robots.
    • Fig. S3. Absorbance spectra of oxyhemoglobin after incubation with commercial purified α-toxin under different conditions.
    • Fig. S4. Absorbance spectra of oxyhemoglobin after incubation with a cocktail of PFTs produced by MRSA bacteria under different conditions.
    • Fig. S5. Characterization of bare Au/Ni/Au NW robots.
    • Fig. S6. Experimental protocol followed to perform the combined application of the RBC-PL-robots: MRSA targeting and PFT neutralization.
    • Legends for movies S1 and S2

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    Other Supplementary Material for this manuscript includes the following:

    • Movie S1 (.mp4 format). Propulsion performance of bare robots in water and whole blood under a US field.
    • Movie S2 (.mp4 format). Propulsion performance of RBC-PL-robots in water and whole blood under a US field.

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