Research ArticleAPPLIED SCIENCES AND ENGINEERING

Organ-on-e-chip: Three-dimensional self-rolled biosensor array for electrical interrogations of human electrogenic spheroids

See allHide authors and affiliations

Science Advances  23 Aug 2019:
Vol. 5, no. 8, eaax0729
DOI: 10.1126/sciadv.aax0729
  • Fig. 1 3D-SR-BAs for electrical interrogation of human electrogenic spheroids.

    (A) 3D-SR-BAs are fabricated using conventional lithography techniques on a sacrificial layer (red arrow). Inset, expanded view of the marked black dashed box of either passive (microelectrode) or active [graphene field-effect transistor (GFET)] biosensors. S and D denote source and drain of the GFET, respectively. (B) Leveraging the prestress in the metal interconnects (gold colored traces), the arrays self-roll upon removal of the sacrificial layer. Inset, expanded view of the marked red dashed box. (C) Cardiac spheroids encapsulated in the 3D-SR-BAs, allowing electrical measurements in 3D. (D) The interface between the cardiac spheroid and array in 2D provides a limited interface for electrical measurement only from the apex of the spheroids.

  • Fig. 2 Highly controlled 3D-SR-BAs.

    (A to C) Bright-field optical microscopy images of photolithographically fabricated 3D-SR-BAs. (A) As fabricated 3D-SR-BA before being released. (B) 3D-SR-BA released with a single turn. (C) 3D-SR-BA released with ~1.7 turns. Scale bars, 100 μm. (D and E) 3D-SR-BAs with varying radii of curvature — simulation and experimental results. (D) 3D-SR-BA with a single turn. (I) A finite element analysis (FEA) simulation result for a 3D-SR-BA with a single turn (inner diameter of ~160 μm). Color bar represents the magnitude of the displacement of the 3D-SR-BA upon removal of the sacrificial layer; SU-8 layers are not shown for visual purpose. (II) A 3D confocal microscopy image of a representative 3D-SR-BA with a single turn (n = 9). (E) 3D-SR-BA with multiple turns. (I) FEA simulation result for a 3D-SR-BA with ~1.7 turns (inner diameter, ~100 μm). Color bar represents the magnitude of the displacement of the 3D-SR-BA upon removal of the sacrificial layer; SU-8 layers are not shown for visual purpose. (II) 3D confocal microscopy image of a representative 3D-SR-BAs with ~1.7 turns (n = 15). Scale bars, 50 μm.

  • Fig. 3 3D-SR-BA with functional passive biosensors (microelectrodes).

    (A) 3D confocal microscopy image of 3D-SR-BA with microelectrodes. Color bar represents the depth in micrometers. Scale bar, 50 μm. (B) Representative cyclic voltammograms (CV) acquired with 1 M KCl at 600 mV/s before (blue trace) and after (red trace) PEDOT:PSS electrodeposition (n = 10). (C) Electrochemical impedance spectroscopy (EIS) plots for the electrodes before (blue trace) and after (red trace) PEDOT: PSS electrodeposition (n = 23).

  • Fig. 4 Effect of 3D-SR-BA on viability of CMs in an encapsulated spheroid.

    (A) Live/Dead assay performed on CM spheroids: a spheroid encapsulated by 3D-SR-BA (top), a spheroid non-encapsulated in 3D-SR-BA (bottom), imaged at (i) 0 hours (immediately after encapsulation), (ii) 1 hour, (iii) 2 hours, and (iv) 3 hours. Green, red, and blue denote live cells, dead cells, and cell nuclei, respectively. Scale bars, 100 μm. (B) Viability analysis of spheroids encapsulated by 3D-SR-BA (blue) and not encapsulated spheroid controls (red) that were imaged every 30 min for 3 hours. Results are reported as mean ± SD (n = 3). There was no significant difference in the % viability between the encapsulated spheroids and the non-encapsulated control spheroids.

  • Fig. 5 Electrical recordings in 3D of cardiac spheroids.

    (A) A 3D confocal microscopy image of 3D cardiac spheroid labeled with Ca2+ indicator dye (Fluo-4, green fluorescence) encapsulated by the 3D-SR-BA. Scale bar, 50 μm. (B) 2D map of the microelectrodes labeled in (A). (C) Representative field potential (FP) traces recorded from the channels labeled in (A) and (B). A.U., arbitrary units. Simultaneously recorded Ca2+ fluorescence intensity as a function of time of the ROI marked by pink box in (A). (D) Averaged FP peak (red trace) and raw data (gray traces, n = 100 peaks recorded by channel 4).

  • Fig. 6 Mapping electrical signal propagation in 3D using 3D-SR-BA.

    (A) Representative recorded single FP fast transients across 12 channels. Red and blue arrows represent the positive and negative phases of the recorded transients, respectively. (B) 2D representation of the fast transient signal phases across all 12 channels. Resting state is presented at t = 14.9 ms, and depolarizing wave propagation is presented at t = 22.0 ms and t = 27.5 ms. (C) 3D rendered signal propagation at t = 22.7 ms. Scale bar, 50 μm. (D) 2D representation of the isochronal map of time latencies. Scale bar, 35 μm. White arrow in (C) and (D) represents average conduction velocity direction.

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/5/8/eaax0729/DC1

    Fig. S1. Schematics of the 3D-SR-BA geometry.

    Fig. S2. Simulation and experimental results for 3D-SR-BA used to estimate the radius of curvature.

    Fig. S3. 3D-SR-BA with microelectrodes: Fabrication and electrochemical characterization.

    Fig. S4. 3D-SR-BA with GFETs: Fabrication and electrical characterization.

    Fig. S5. Electrical recordings with and without addition of chemomechanical decoupler, blebbistatin.

    Fig. S6. Simultaneous Ca2+ and electrical recording.

    Fig. S7. Several hours (3 hours) 3D recording.

    Fig. S8. 3D electrical signal propagation in spheroids.

    Table S1. Thickness and mechanical parameters of SU-8, Cr, and Pd used in FEM simulations.

    Table S2. Data summary for the Raman analysis of LPCVD synthesized graphene before and after patterning.

    Movie S1. Release of the 3D-SR-BA upon sacrificial layer etch.

    Movie S2. Unrolling the 3D-SR-BA using a micromanipulator.

    Movie S3. Beating CM spheroid stained with Ca2+ (Fluo-4) encapsulated in the 3D-SR-BA (associated Ca2+ imaging analysis is presented in fig. S6).

    Movie S4. Electrical signal propagation in CM spheroid presented in Figs. 4 and 5.

  • Supplementary Materials

    The PDF file includes:

    • Fig. S1. Schematics of the 3D-SR-BA geometry.
    • Fig. S2. Simulation and experimental results for 3D-SR-BA used to estimate the radius of curvature.
    • Fig. S3. 3D-SR-BA with microelectrodes: Fabrication and electrochemical characterization.
    • Fig. S4. 3D-SR-BA with GFETs: Fabrication and electrical characterization.
    • Fig. S5. Electrical recordings with and without addition of chemomechanical decoupler, blebbistatin.
    • Fig. S6. Simultaneous Ca2+ and electrical recording.
    • Fig. S7. Several hours (3 hours) 3D recording.
    • Fig. S8. 3D electrical signal propagation in spheroids.
    • Table S1. Thickness and mechanical parameters of SU-8, Cr, and Pd used in FEM simulations.
    • Table S2. Data summary for the Raman analysis of LPCVD synthesized graphene before and after patterning.
    • Legends for movies S1 to S4

    Download PDF

    Other Supplementary Material for this manuscript includes the following:

    • Movie S1 (.mp4 format). Release of the 3D-SR-BA upon sacrificial layer etch.
    • Movie S2 (.mp4 format). Unrolling the 3D-SR-BA using a micromanipulator.
    • Movie S3 (.mp4 format). Beating CM spheroid stained with Ca2+ (Fluo-4) encapsulated in the 3D-SR-BA (associated Ca2+ imaging analysis is presented in fig. S6).
    • Movie S4 (.mp4 format). Electrical signal propagation in CM spheroid presented in Figs. 4 and 5.

    Files in this Data Supplement:

Stay Connected to Science Advances

Navigate This Article