A PARYLENE PERIPHERAL NERVE CUFF ELECTRODE WITH INTEGRATEDMICROFLUIDICS
A. Cobo1, K. Scholten1, J. Yoo1, C. Larson1, T. Hudson1, V. Pikov2, and E. Meng1*
1
University of Southern California, Los Angeles, California, USA
2
GlaxoSmithKline, UK

ABSTRACT
We present a novel micromachined polymer peripheral nerve
interface that combines both electrodes and microfluidic channels inan adjustable cuff sized to interface with rat sciatic nerve. Parylene
fluidic channels enable targeted delivery of lysing agents andneurotrophic factors to fascicle sites on the nerve surface, in order
to induce axonal sprouting towards electrodes. This Lyse-andAttract Cuff Electrode (LACE) uses a simple locking mechanismthat is adjustable for close contact with nerves of varying diameters.
We demonstrate fully functional cuffs having electrode impedancesof < 2 kΩ (1 kHz) and controlled microfluidic infusion at < µL/minflow rates.

INTRODUCTION
Neural recording and stimulation technologies are needed tointerface with peripheral nervous system (PNS) to create abidirectional link between the nervous system and artificial devices(prosthesis) for restoration of sensorimotor function (Figure 1) [14]. Electrical interfaces to the peripheral nerves are either
extraneural or intrafascicular/intraneural. Minimally invasive
extraneural interfaces are placed on the circumference of the nerve
which is surrounded by a collagenous sheath (epineurium) that
limits access to individual nerve fibers. Improved access toindividual fascicles has been achieved by extraneural interfaces that
reshape the nerve by flattening it, as is the case of the previously
described flat interface nerve electrode (FINE) [5]. However thesignal to noise ratio is still restricted by the impedance of theepineurium, and nerve damage can be induced by reshaping.
Intrafascicular/intraneural electrodes gain greater access toindividual nerve fibers, improving signal-to-noise ratio, and canselectively target small groups of axons, but are significantly moreinvasive [6, 7]. Electrodes may either be inserted longitudinally into
the nerve and run parallel to nerve fibers, or transversely and run
perpendicular to fibers. Of the two methods, transverse electrodes
are associated with greater risk of nerve damage. Invasively
inserted electrodes are fabricated from materials with elastic moduli
several orders of magnitude greater than that of nerve, contributing
to chronic tissue response and the degraded recording/stimulation
stability over time.
Neither approach has been able to achieve the desired
selectivity while minimizing invasiveness, and all prior attempts
suffered from a lack of chronic reliability. To overcome thesehurdles and achieve a long-term stable interface with high fidelity
recordings, we developed a non-invasive approach in which a
polymer cuff electrode targets individual fascicles within a nerve by
inducing the sprouting of axonal collaterals towards electrodes
embedded in the microfluidic channels of the cuff. Axonal sprouting
will be selectively induced from the fascicles closest to themicrofluidic outlet ports via localized delivery of lysing agents to
temporarily disrupt the epineurium, followed by delivery ofneurotrophic factors to promote axonal sprouting towards electrodes
within the microchannels [8-11]. The design calls for a cuff withadjustable diameter to ensure localized diffusion of chemicals into
the epineurium of nerves of varying diameters. This Lyse-andAttract Cuff Electrode (LACE) aims to increase fascicular
selectivity and signal-to-noise ratio for recording and/or stimulation

without traumatic invasiveness, while providing long-term stability
beyond the current state of the art.

Figure 1: Peripheral nerve interfaces necessary for braincontrolled prostheses.

DESIGNThe LACE is fabricated from thin-film biocompatible polymer
Parylene C (Figure 2), and incorporates 4 surface micromachined
microfluidic channels (250 µm W × 20 µm H) for targeting four
fascicles in a rat sciatic nerve. Each microfluidic channel contains apair of Pt electrodes (300 µm W × 1500 µm L) for differential
recording and/or stimulation. The cuff wraps around a nerve withthe microfluidic outlets in direct contact with the circumference ofthe epineurium, and is held in position by etched serrations (Figure2). The cuff is adjustable (1.1-1.5 mm diameter, 0.1 mm increments)
to provide close fits to nerves of varying diameter. Ancillarycomponents identified in Figure 2b include a port for external fluidic
connection, Parylene ribbon cable (~2 cm long) for external
electrical connection, and a reference electrode external to themicrofluidic channels.

Figure 2: (a) Schematic of LACE for targeting individual fascicles
within a nerve. Insert shows the cross-sectional view. (b) Fabricated
LACE in which microfluidic channels are highlighted by thepresence of photoresist.
Fabrication
Devices were fabricated on a silicon wafer for mechanicalsupport during the microfabrication process (Figure 3). A 10 µm
Parylene base layer was first deposited by chemical vapor
deposition. Sputtered Pt electrodes and leads (2000 Å) were thenpatterned by a liftoff process with negative photoresist AZ 5214 EIR. A 10 µm Parylene insulation layer was then deposited andselectively plasma-etched to expose electrodes and contact pads.
Sacrificial photoresist AZ 4620 was spun to a height of 20 µm andpatterned to form the microfluidic channels. An 8 µm Parylene layerwas then deposited over the photoresist. Oxygen plasma etchingcreated openings for microfluidic ports as well as the device cutout.
Devices were released from the substrate and sacrificial photoresist
was removed with an acetone soak. No stiction of Parylene
structures was observed.

Figure 5: Prototype LACE interlocking mechanism test. (a) Flat
LACE, (b-c) tab threaded through first slit and kept in place byserrated teeth, and (d) tab secured by threading through second
slit to form the buckle locked structure.
Figure 3: LACE fabrication process which utilizes standard surface
micromachining techniques for Parylene.
Device level fluidic packaging consisted of a custom madepolyurethane (PU) catheter (SAI Infusion Technologies) secured inplace with MED 4210 silicone adhesive (Factor II Incorporated). A
zero-insertion-force connector (ZIF, part no. 0514411072 Molex
Inc) was used to establish reversible electrical connections to thecuff electrodes through the contact pads at the end of the integrated
Parylene ribbon cable (Figure 4). The ZIF connector was mated to aflat flexible cable (FFC, part no. 050R10-102B, Parlex USA LLC)
and a rigid PCB for external connection to the electrophysiologicalrecording setup.

Figure 4: Cuff electrode with integrated Parylene cable is attached
to a zero insertion force (ZIF) connector.

METHODSInterlocking Mechanism
The cuff structure consisted of an elongated tapered tab thatloops around the sciatic nerve, inserts into an etched slit and locks
in place using etched serrated teeth (Figure 5a-c). A two-partlocking mechanism was used to maintain the loop around the nerve,
and includes an additional slit through which the tab is threaded tocreate a secure buckle structure (Figure 5d). The guide needle hole
was utilized to pass a suture from the circular hole in the tab andfacilitate guiding of the tab into the slits. Devices were wrapped andunwrapped around simulated agarose nerve phantoms and holding
strength was evaluated under gentle handling conditions (n > 10).
An agarose concentration of 1.37 % wt/wt in water was selected
since it has a compressive modulus equal to that of rabbit sciatic
nerve [12, 13].

Microfluidic Channel Evaluation
Evaluation of the microfluidic channels and fluidic connection
was conducted by driving colored dye through the channels using
an infusion pump (Harvard apparatus Infuse/Withdraw PHD 2000)
and imaging the progression of the dye with a microscope (Caltex
Scientific HD60T). The LACE device was submerged in distilledwater under the microscope, and primed by flushing the channels
with isopropyl alcohol and distilled water for 5 minutes each. With
the LACE device in a flat or curled orientation, a solution of dilute
green dye was infused into the LACE device at varied flow rates
ranging from 5 to 2000 nL/min.
Maximum possible flow rate of the LACE device wasdetermined by pumping dye through a LACE device wrappedaround a nerve phantom, and observing which flow rates induced
leaks or structural failure. Flow rates were increased from 300
nL/min at increments of 100 nL/minute. Between each increment,
the device was uncurled and infused with green dye in a flatorientation under the microscope in order to detect any small leaks
that may have formed in the interior of the channels, then re-curled
for the next flow rate.
Localization of fluid delivery was demonstrated by locking a
LACE device around a simulated sciatic nerve phantom, thendriving sodium fluorescein dye (Sigma F6377) through themicrofluidic channels using the infusion method previously
described at a flow rate of 50 nL/min.
Electrode Evaluation
Cyclic voltammetry (CV) and electrochemical impedance
spectroscopy (EIS) were used to assess the stimulation andrecording capabilities respectively of the cuff electrodes.
Electrochemical characterization of the electrodes was performed
with a Gamry Reference 600 potentiostat (Gamry Instruments,
Warminster, PA). Prior to EIS measurement, CV was used toelectrochemically clean the electrode surface and take initial
measurements. LACE devices were immersed in 0.05 M H2SO4
with a constant stream of N2 in a three-electrode cell. The working
electrode was cycled between −0.2 to 1.2 V with respect to an
Ag/AgCl (3M NaCl) reference, a potential range that is within thewater window [14]. A 1 cm2 Pt plate served as a counter electrode.
Using a scan rate of 250 mV/s, each electrode was cycled for 30
cycles. EIS was performed in phosphate buffer saline (1× PBS) atroom temperature with an AC perturbation signal of 10 mV (rms) in
the frequency range of 1-105 Hz. As with CV, an Ag/AgCl (3M
NaCl) reference and 1 cm2 Pt plate counter were used.

RESULTS
Interlocking Mechanism
The locking mechanism was simple to implement and robust.
A suture needle facilitated wrapping the device around an agarose
nerve phantom of 1.2 mm diameter. The LACE was able to wrap
tightly around the phantom without sliding of the device or damage
to the simulated nerve. Once locked in place, the LACE did notdisengage during fluidic and electrical testing, evidence of a securelock. No damage to the cuff was observed despite repeatedly
engaging and releasing the lock and curling and uncurling the LACE
around the nerve phantom.
Microfluidic Channel Evaluation
No leakage or delamination was observed up to a flow rate of2000 nL/min, with uniform flow through all fluidic channels andoutlet ports in the flat configuration (n = 4). Figure 6a displays asequence of images showing simultaneous and equally distributed
flow out of all four outlet ports in the flat orientation. Image analysis
confirmed that flow rate through channels was uniform within 6%
standard error. In the curled configuration, flow was achieved in 3
out of 4 channels which is sufficient for the in vivo application(Figure 6b). Flow through the distal channel was blocked bypinching of the horizontal feeder channel and can be mitigated bychannel design changes such as: reduced feeder channel width andincorporation of Parylene support walls along the horizontal channel[15, 16]. These changes have been incorporated and are being
evaluated.

electrodes can be characterized by their cathodal charge storage
capacity (CSC), which is calculated from the time integral of thecathodic current in a cyclic voltammogram per unit geometricsurface area (GSA) [19] as shown in equation (2).
!"# =

&'
210 +,-./

(1)

,", =

&-0123456"#(2)

The cyclic voltammogram obtained for all electrodes tested
indicated a clean electrode surface. The ESA calculated was
635,714 µm2, which is different from the GSA of 450,000 µm2 due
to surface topology. The CSC was 1.06 ± 0.06 mC/ cm2 and theroughness factor, defined as the ratio of ESA to GSA was 1.4 (n =6). No bubble generation was observed during the CV, which isimportant for safe electrical stimulation.

Figure 7: 1.37% (w/w) agarose nerve phantom after localization
testing with fluorescein dye.

Figure 6: Sequential photographs of the infusion experiment at 830
nL/min flow rate in a (a) flat device and (b) curled device. Image
analysis using ImageJ software confirmed that flow rate throughchannels was uniform within 6% standard error.
During maximum flow rate testing, microfluidic channel inlet
failure occurred at a rate of 1000 nL/min. The leak appeared to befrom delamination between Parylene layers. A safe flow rate of 900
nL/min can be achieved by the LACE device, which greatly exceeds
the typical epineural space drug delivery rate of 17 nL/min that will
be employed in our studies [17]. Localized drug delivery wasachieved with the LACE as shown in Figure 7. In on-going testing
the agarose nerve will then be imaged using a fluorescent
microscope (Nikon LV100; mercury 100 W lamp, Chiu Technical
Corporation; B-2A filter cube) and digitally processed in ImageJ inorder to correlate fluorescent signal with dye concentration.
Electrode Evaluation
The expected standard voltammogram for Pt immersed insulfuric acid was achieved for all electrodes. Distinct peaks ofcurrent in the cyclic voltammogram result from the process ofhydrogen atoms adsorption to and desorption from the electrodesurface were observed as shown in Figure 8. These peaks can be
integrated with respect to the scan rate to calculate the electroactive
surface area (ESA). In equation (1), QH is the hydrogen desorption
charged measured via CV and 210 µC/cm2 is the value for thecharacteristic charge density associated with the monolayer ofhydrogen atoms adsorbed to polycrystalline Pt [18]. Stimulation

Figure 8: Cyclic voltammetry in H2SO4 of single recording siteyielded standard Pt electrode characteristics.
EIS measurements on cuff electrodes yielded an electrical
impedance of 1.8 ± 0.11 kΩ and a phase of -56.4 ± 1.0º at 1 kHz
demonstrating low variation between electrodes (n = 6). A
representative EIS measurement for a single electrode site post CVclean is shown in Figure 9. Recorded impedances are expected toprovide adequate neural recording potential. Baseline EC
measurements on cuff electrodes demonstrated suitable electrodeproperties for peripheral nerve recordings and stimulation.

CONCLUSION
We developed a peripheral nerve interface that combines bothelectrodes and microfluidic channels in an adjustable cuff sized tointerface with rat sciatic nerve. The LACE device uses a robust andsimple locking mechanism that is adjustable for close contact withnerves of varying diameters. Parylene C was selected as thestructural material for its biocompatibility and improved mechanicalmatching to tissue.
We demonstrated fully functional cuffs having low electrode
impedances of < 2 kΩ (1 kHz) to help reduce susceptibility to noise
pickup from sources such as muscle outside the cuff or recording
equipment [20]. Controlled microfluidic infusion at < µL/min flow
rates and localized drug delivery was achieved, which is importantto generate specific drug gradients necessary for guiding the axons
into the microchannels. Future work entails characterization ofmodified fluidic channels to allow uniform flow through allchannels in the curled orientation in order to target each fascicle
inside the sciatic nerve, long-term characterization of electrodes,
and preparation for in vivo animal testing.

[6]

[7]

[8]

[9]

[10]

[11]

[12]
Figure 9: EIS in PBS of single recording site showed a low
impedance (< 2 kΩ) at 1 kHz.

[13]

ACKNOWLEDGEMENTS
This work was supported under DARPA cooperative
agreement HR0011-15-2-0006, Viterbi School of Engineering Ph.
D. Merit Fellowship (AC), and USC Provost Fellowships (JY and
TH). The authors would like to thank Dr. Donghai Zhu and themembers of the USC Biomedical Microsystems Laboratory for theirassistance with this project. Travel support was provided by OAI.
An OAI hybralign series 200 mask aligner was used for processingof Parylene devices.

[14]

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CONTACT*E. Meng, tel: +1-213-821-3949; ellis.meng@usc.edu