Available online at www.sciencedirect.
com
Status and challenges of electrical stimulation use in
chronic wound healing
Miruna Verdes 1,2, Kimberly Mace 3, Lee Margetts 4 and
Sarah Cartmell 1,2
Non-healing wounds have led to a soaring clinical and
socioeconomical need for advanced wound-care techniques.
Electrical stimulation is an emerging therapy inspired by the
wound's endogenous electric field. Promising results of clinical
trials have encouraged efforts to create wearable stimulation
devices, uncover multiple cellular targets, and optimize
stimulation regimes. However, the field faces a translational
bottleneck. This review aims to highlight the gaps between in
vivo treatments and in vitro associated experiments by
discussing the current knowledge of the generation,
characterization, and targets of electrical stimuli. It becomes
clear that enabling the translation of this technology will require
increasing the complexity of the current models for skin
endogenous and controlled ion transport, and investigating
which stimulus has an optimum effect on cells derived from
chronic wound-prone patients.
Addresses
1
Department of Materials, Faculty of Science and Engineering, The
University of Manchester, Manchester M13 9PL, UK
2 surface [2]. While the structure of the epidermis mostly
The Henry Royce Institute, Royce Hub Building, The University of
Manchester, Manchester M13 9PL, UK
3
Division of Cell Matrix Biology and Regenerative Medicine, Faculty of
Biology, Medicine and Health, University of Manchester, Manchester
M13 9PT, UK
4
Department of Mechanical, Aerospace and Civil Engineering, Faculty
of Science and Engineering, The University of Manchester, Manchester
M13 9PL, UK
Corresponding author:
Sarah Cartmell (sarah.cartmell@manchester.ac.uk).
Current Opinion in Biotechnology 2022, 75:102710
This review comes from a themed issue on Tissue, cell and
pathway engineering
Edited by Lorenzo Moroni and Martijn van Griensven
For complete overview of the section, please refer to the article
collection, “Tissue, Cell and Pathway Engineering”
Available online 7th April 2022
https://doi.org/10.1016/j.copbio.2022.102710
Crown Copyright © 2022 Published by Elsevier Ltd. All rights
reserved.
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]]
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Introduction
Skin is the largest human organ. Its main function is to
help our bodies maintain homeostasis by acting as a
selective barrier between the inside and the outside,
offering protection from pathogens, and damaging phy­
sical and chemical agents, while also regulating the
movement of heat, water, and electrolytes across it. This
filter function is conveyed by a complex layered struc­
ture of interdependent cells and the matrix surrounding
them, with the highest adaptability and turnover being
characteristic of the two outermost layers, epidermis and
dermis [1]. These are two contrasting layers, connected
through a highly specialized extracellular matrix (ECM)
called the basement membrane. The epidermis relies on
the dermis for both mechanical stability and nutrition.
Both are enhanced by the interdigitating pattern of
epidermal downgrowths (rete ridges) within the upper
dermis and dermal protrusions (papillae) within the
epidermis, which give an overall increased junction
consists of tightly interconnected cells, the dermis has a
highly fibrillary architecture, within which cells are
sparsely distributed [3]. Concurrently, blood vessels are
contained within the dermis, and nutrients must diffuse
from the dermal capillary networks, through nanometer-
sized pores in the basement membrane, to the epi­
dermis [2].
Healthy skin, as do all the organs in our body, maintains
an electric potential difference across its epithelium.
Epithelial cells in the skin, called keratinocytes, have a
heterogeneous distribution of ion channels. Ions are ac­
tively transported across the cell membrane against their
electrochemical gradient to generate and preserve a
transepidermal potential ranging from 15 to 60 mV [4],
with a positive charge inside (Figure 1a). Concomitantly,
some of the positively charged ions flow back to the
apical side of the epithelium through the paracellular
pathway to re-establish the electrochemical equilibrium.
However, this is limited by the tight cellular junctions,
which translate into a high electrical resistivity [4].
Hence, while the epidermis is intact, the transepidermal
potential is maintained [4,5]. It has been suggested that
sodium and potassium are the main contributors to this
circuit [4], but this information is yet to be confirmed in
the skin. Conversely, measurements with ion-selective
probes conducted in corneal epithelium wounds showed
Current Opinion in Biotechnology 75(2022) 102710
2 Tissue, cell and pathway engineering

Figure 1

Current Opinion in Biotechnology

The skin electric field. (a) Endogenous electric potential in the intact (left) and wounded (right) skin. What we know is that keratinocytes pump ions to
maintain a transepidermal voltage gradient of 20–50 mV. Wounds are low-resistance paths through which the positive ions accumulated underneath
the epidermis can escape to the surface and generate a lateral EF. What we do not know is whether there is also a voltage gradient across the dermis
(1), and therefore do not know the input signal affecting fibroblasts, macrophages, and endothelial cells. As the layer of keratinocytes actively pumps
ions, the dermoepidermal junction could bear a positive charge relative to the deep reticular dermis. If this is true, in full-thickness wounds, there
should also be an inward positive ion flow, from the dermoepidermal junction towards the hypodermis. The newly available positive ions in the deeper
dermis (2) can now flow along the collagen fibers, and with a higher speed along the coaxial fibers in the reticular dermis, making the wound bed the
‘anode of the wound’ in comparison to the surrounding dermis. This charge transport path could in turn naturally stimulate the electrotaxis of
fibroblasts and macrophages towards the wound bed. (b) By contrast, we look at the effects of an exogenous stimulus, such as the one generated by
a wound dressing, with a central cathode and the anode on the surrounding intact skin. We predict that placing the cathode on the wound bed of a full-
thickness wound could cancel out the transepidermal potential forcing keratinocytes to pump faster (3), and reverse the ion flows (and implicitly the
cellular stimulus) both on the skin surface (4) and within the dermis (5). Here, charge density is a measure of the number of positive charges within a
unit volume.

Current Opinion in Biotechnology 75(2022) 102710 www.sciencedirect.com

 Reviewing chronic wound bioelectricity principles Verdes et al.
that chloride, calcium, and potassium ions are actually
the main contributors to the naturally occurring outward
current [6].
Damage to the skin is not uncommon, and complex
processes ensure that the skin barrier is restored in a
timely fashion. Our cells have evolved to recognize the
breach and synchronize with their neighbors to stop the
bleeding, clean up damaged tissue, and generate new
dermal ECM on top of which keratinocytes can migrate
and close the wound before pathogens can become es­
tablished. After re-epithelialization, in the absence of
imminent pathogen threats, the mechanical properties of
the skin are restored through a long process of re­
modeling [7,8].
However, this process can often fail, and non-healing
wounds are a growing societal burden. In the UK alone,
the annual prevalence of wounds increased by 71% be­
tween 2012 and 2013 and 2017 and 2018, and 67% of the
£8.3 billion budget allocated for wound care in the latter
period was spent on wounds that did not heal within that
year [9]. Several intrinsic factors make the skin less re­
sistant to mechanical stress, and impede healing, as most
chronic wounds are either pressure, diabetic, or venous
skin ulcers [10]. These comorbidities may lead to a
suboptimal ECM structure [11,12] and cellular behavior
[13–20], which can then contribute to thinning of the
epidermis, reduced skin elasticity, tissue necrosis, and
eventually wounds [11]. In addition, when the epidermal
barrier is breached for a long time, wound beds are
further disturbed by bacterial colonies forming biofilms
[21]. The wound then fails to exit the inflammatory
phase, whereby the ECM is excessively degraded, and
the wound cells become senescent and less responsive to
stimuli [7,22].
Acute skin wounds generate a lateral electric field (EF)
of about 100–200 V/m, with the wound edge being more
positive than the center [4,23], as ions escape through
the epidermal breach and higher current densities are
recorded on the wound outline (Figure 1a). This, cou­
pled with the observed keratinocyte migration towards
the cathode of an EF, encouraged the development of
electroactive wound dressings in which the cathode is
connected to the wound bed and the anode to the edges
and surrounding skin (Figure 1b). Measurements in
corneal wounds of diabetic mice suggest that the EF is
indeed weaker in some chronic wounds [24]. However,
measurements of the endogenous EF in chronic wounds
remain a fundamental gap in the literature.
Although electrical stimulation (ES) is a versatile tool for
wound healing, devices and treatment regimens still
need to be optimized for this therapy to reach its full
potential [25,26]. An effective chronic wound treatment
should target ischemia by enhancing nutrient-rich blood
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3
flow into the wound, inhibiting biofilm formation, and
finally directly targeting the cells to encourage a pro-
healing behavior [21]. To date, ES has been shown to
promote angiogenesis [27–29], disrupt biofilms [30], and
induce various responses in the cells involved in wound
healing (Tables 1 and 2). These results should en­
courage further exploration, as different stimulation
techniques may lead to better results. Here, the pitfalls
and challenges of using ES to target the different cellular
agents of wound healing will be discussed.
The signal
The EF and the resulting endogenous ionic currents are
one of multiple wound-healing guiding cues. Changes in
the local skin EF appear immediately upon wounding,
and are gradually dampened as the wound heals and the
epidermal resistance is recovered [4]. Cells that play a
key role in wound healing, such as keratinocytes [31],
macrophages [32], fibroblasts [33], and endothelial cells
[34,35], were shown to react to such changes.
Aiming to reproduce the healing benefits of the en­
dogenous EF, we developed a range of ES devices for
the in vivo stimulation of chronic wounds [26]. The ad­
vantage of these over other guiding cues, such as damage
or pathogen-associated molecular patterns, or various cell
signaling molecules is that exogenous ES can in theory
be more precisely controlled. The current ES devices
consist of electrodes in direct contact with the skin
[26,36], with the option of adding state-of-the-art tech­
nology such as nanogenerators, nanostructured con­
ductive dressings, or nanoparticles with ES-dependent
drug release to improve the wearability, compatibility,
and functionality [37,38]. The cathode is generally
placed in the middle of the wound, while the anode
either surrounds the wound edges or is placed on one
side of the wound [26].
Although any electric potential gradient can be gener­
ated with adequate electric devices, we are still limited
in controlling the cellular input in vivo by the scarce
knowledge regarding the electrical properties of the
transduction path. Firstly, even though the electrical
properties of the skin are sensitive to physiological
changes and vary significantly between the body loca­
tions of the same individual [39,40], to date there is no
study that has measured the electrical properties in pa­
tients with chronic ulcers, or in locations prone to such
wounds (sacral region or legs). Secondly, the only in vivo
skin EF sensors, the vibrating probe [41] and the Der­
macorder [23], can only measure the EF at the wound
surface. While these are useful in determining the EF
impacting the keratinocytes at the wound edge and
bacteria settling on the wound surface, they do not
provide information about the EF of the deeper dermis.
This information is important, as current electroceuticals
Current Opinion in Biotechnology 75(2022) 102710
4 Tissue, cell and pathway engineering

may in fact dampen or even reverse the directional

Reference
electric cue arriving at fibroblasts and macrophages re­
[64] sident in the dermis surrounding the wound (Figure 1b).

[67]

[66]

[27]

[65]

[33]

[70]
constant, polymerized actin, and podosomes polarized toward the
In silico models can predict the endogenous EF [42] or

Anode (preceded by elongated morphology perpendicular to EF)

Anode (preceded by elongated morphology with total cell area

the impact of electrodes applied to wounded skin within

the deeper layers [43,44]. However, there is still space
for improvement, as these models do not account for the
Perpendicular to EF (laminin and fibronectin substrate)

ability of keratinocytes to maintain the transepidermal

potential gradient, a physiological charge distribution
within the dermis, and assume that the skin layers are
isotropic materials when this may not be the case [45]. In
silico simulations of charge transport combined with
Anode (peaking at 4 hours of ES)
high-precision ECM images [46] should now constitute a
viable solution for building a more representative model
leading edge, facing the anode)

of ion conduction within and around wounds. Validation

Cathode (Ca2+ independent)
Random (glass substrate)

of these models may be done using the methods for

Avg speed: 0.05 µm/min

Avg speed: 0.15 µm/min

Enhanced (Scratch test)

studying the endogenous ion distribution within the
Direction of migration

Random enhanced

skin [47].
DC 1 hour, 100 V/m Not significant

Various in vitro cellular ES techniques have also been

DC, 2 hours 150 V/m Cathode

developed. Classing by the EF generation method,

DC 1 hour, 400 V/m Anode

these involve either direct, capacitive, inductive cou­

pling, or stimulation via conductive scaffolds [36,48,49].
While in vitro setups offer better control over the ex­
DC up to 6 hours,
AC, 1 Hz, 90 min,

perimental conditions, the complexity of the cell culture

medium still makes the characterization of the cellular
DC, 24 hours,

DC > 1 hour,
100–400 V/m

150–200 V/m

25–100 V/m
DC, 10 min,
Electrotaxis of monocytes, macrophages and fibroblasts in response to electrical stimulation.

DC, 1 hour,

electric microenvironment challenging. Therefore, it is

200 V/m

100 V/m

200 V/m

still difficult to compare the results from different in vitro

methods on one hand, and unclear how well they model
existent electroceuticals’ effect on the skin in vivo on the
other hand. In silico models can also assist the char­
Ag/AgCl electrodes and calf

Conductive PPy/HE/PLLA

acterization of cellular electric microenvironments in

Stimulus

vitro, but their limitations must not be forgotten.

Platinum electrodes
Direct coupling Ag/AgCl electrodes

Ag/AgCl electrodes

collagen I coating

For the direct coupling setup, a common simple ap­

proximation of the stimulus is the ratio between the
membranes

voltage gradient applied on the electrodes and the dis­

tance between them. At the same time, the generated
stimulus can also be spatially resolved at different levels
of precision using commercially available software
packages that are used to model electrolytic cells
ES strategy

ES through

[50–52]. Contrary to what the first approximation im­

scaffold

plies, the in silico simulations show that the generated

EF is not homogenous, leading to higher current den­
sities near the electrodes [50,51]. In vitro, this translates
Bovine pulmonary artery
Fibroblasts Murine NIH 3T3&SV101

into cell death or different cellular responses throughout

the culture dish, showing the need to delimit the culture
Human dermal

area [50]. In vivo, this should argue for developing me­

(embryonic)

chanisms that can ensure that currents in all wound areas

Human macrophages

are maintained underneath a biocompatible threshold.

Human monocyte

While current in silico models have proved useful in this

case, they are still an approximation, as the culture
medium is a concentrated electrolyte with multiple
Table 1

charge carriers and macromolecules that may impact the

Cell

transduction of time-dependent ES.

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 Reviewing chronic wound bioelectricity principles Verdes et al. 5

Table 2
Functionality of macrophages and fibroblasts harnessed through electrical stimulation.

Cell ES strategy Stimulus Cell response Ref

Human Direct coupling with No extra stimulus DC, 2 hours, 150 V/m Phagocytosis enhanced [64]
macrophage Ag/AgCl electrodes After 24 hours:
TNF-α: moderate increase (M1-like)
NT3: significant increase (M2-like)
CCL2, IL10, VEGF, TGF-β: constant
PI3K and p-ERK activated
IFN-γ and LPS (M1 IL1b increased
stimuli) IL-23 increased
IL-6 constant
IL-10 constant
TNF-a constant
IL-4 (M2 stimulus) TNF-α enhanced
VEGF constant
IL-10 constant
TGF-b constant
Human dermal PCL scaffold with EF Step pulses, 3 hours, α-SMA 12.4× higher expression [71]
fibroblast aligned uniaxial fibers 50 V/m FGF2, EGF, FN secretion elevated
and TGF-β1 (compared with just TGF-β1 and scaffold)
ES through scaffold Conductive PPy/HE/ DC, 6 hours, 200 V/m Contractility enhanced [70]
PLLA membranes α-SMA expression enhanced (starting at
2 hours ES)
FGF1, FGF2 secretion enhanced
DC, 6 hours, Production increased: CCL7, KGF, TIMP2 [69]
50–200 V/m Production decreased: MMP2
Ppy-coated fabrics Step pulses, 24 hours, TGF-β1, p-ERK, NF-kB upregulated [72]
100 V/m Changes maintained in subculture and in
vivo transplants
Direct coupling Carbon rod electrodes Step pulses, Collagen, MMP1, elastin significant [73]
Confluence ≥ 90% 0.5–24 hours, expression increase at applied EF above
0–1000 V/m 300 V/m, and stimulation longer than
2 hours

In other in vitro models, electrolysis is avoided to pre­
vent generating harmful byproducts [49], but stimulus
characterization becomes more challenging. Briefly, cell
culture wells can be placed between the plates of a ca­
pacitor (capacitive coupling), near an inductor coil (in­
ductive coupling), or on top of a biocompatible
conductive scaffold. While the interaction of capacitor
plates with dielectric materials and inductor coils with
conductors is well understood, new models are required
to describe their interaction with electrolytes within the
cell culture recipients. Similarly, the principles of in­
teraction of conductive polymers with biological cells
and their electrolytic culture medium are yet to be de­
scribed and modeled. As such, even though electricity is
involved in all those in vitro setups, it is not clear whe­
ther they all generate the same cellular stimulus, how to
compare them, or how well they are translated into
current electroceuticals.
The targets
The first studies to suggest that ES can promote wound
healing were focused on epithelial cells, such as kerati­
nocytes in the skin [4]. Several studies have shown that
keratinocytes migrate toward the cathode of an EF
[53–55]. In chronic wounds, even though keratinocytes
are highly proliferative at the wound edges, they fail to
migrate over the wound bed and seal the lesion [56].
While ES is supposed to override this problem, in vitro,
keratinocyte electrotaxis is blocked by the absence of
extracellular calcium [57], and, additionally, the direc­
tion can be reversed by lowering the extracellular pH
[58]. Conversely, as the wound heals, the pH moves
from neutral to acidic, and wounds with a higher pH are
shown to heal slower [59,60]. Moreover, even if the
surface pH is right, keratinocyte electrotaxis may still be
inhibited by inflammatory cytokines in the wound bed,
or, in full-thickness wounds where the dermal layer is
damaged, by an incomplete granulation tissue that they
cannot connect with [61]. These are both problems that
could be introduced by suboptimal macrophage and fi­
broblast behavior in chronic wounds.
Successful wound healing and re-epithelialization are
dependent on the dynamic evolution of the in­
flammatory phase and transition to the repair phase, a
process tightly regulated by macrophages and fibroblasts
[8,62]. The former is characterized by the majority of
macrophages exhibiting a pro-inflammatory phenotype
that promotes host defense and removal of damaged
tissue while fibroblasts are recruited to the provisional

www.sciencedirect.com Current Opinion in Biotechnology 75(2022) 102710

6 Tissue, cell and pathway engineering
Table 3
Cellular electrical properties and their fast response to ES.
Cell Device Stimulus
Macrophage Human Glass None
microelectrode Depolarizing current
12.7–30.5 V/s
Direct coupling DC, 2 hours 150 V/m
Ag/AgCl electrodes
Murine Patch clamp Voltage pulses and ramp
cardiac −100 to +60 mV
Human fibroblast Direct coupling 1 Hz; 30 min, 200 V/m
Platinum electrodes DC; short term, 1000 V/m
AC; 1 Hz; short term,
1000 V/m
AC; 1 Hz; 30 min, 200 V/m
AC; 10 Hz; 30 min,
200 V/m
AC; 100 Hz; 30 min,
200 V/m
ECM wound bed. Subsequently, within the latter phase,
the pro-repair macrophages become the predominant
phenotype. At the same time, the recruited fibroblasts
degrade the provisional ECM fibrin, replace it with
collagen-rich granulation tissue, convert it to myofibro­
blasts, and contract the wound [63]. Both macrophages
and fibroblasts secrete several other growth factors,
which can, in turn, regulate re-epithelialization and an­
giogenesis by their impact on keratinocytes and en­
dothelial cells. Therefore, only these two will be further
focused on as ES targets in chronic wound healing.
In vitro experiments show that recruitment of both
macrophages and fibroblasts to the wound bed could be
targeted with direct current (DC) ES of physiological
strength (Table 1). However, they require a longer and
opposite stimulus to keratinocytes, a fact that might not
be addressed by some in vivo treatments and wound
dressings (Figure 1b). Both macrophages and fibroblasts
exhibit electrotaxis toward the anode of a DC EF in a
voltage-dependent manner [33,64], and require more
than an hour of ES to begin migrating directionally
[33,64,65]. Moreover, establishing the origin and differ­
entiation status of the targeted cells will play an im­
portant role in the in vivo chronic wound therapy results.
In vitro, while mature macrophages migrate toward the
anode [64], their bone marrow-derived progenitors,
monocytes, are directed toward the cathode [64]. Simi­
larly, fibroblasts from embryonic-origin cell lines were
also reported to migrate toward the cathode [66]. On top
of this, the way in which the tissue transduces the exo­
genous stimulus may also impact the recruitment result.
Mature macrophages exposed to an alternating current
EF migrate perpendicular to the field direction [67].
Current Opinion in Biotechnology 75(2022) 102710
Cell response Reference
Resting membrane potential [−49, −72] mV [83]
Repetitive Ca2+-dependent spikes [83,84]
Gradual rise in intracellular Ca2+ peaking at 2.4 min [64]
of ES
A computational model for the macrophage's [82]
electrophysiological properties was developed.
Reported properties are:
▪ membrane resistance 2.2 ± 0.1 GΩ
▪ capacitance 18.3 ± 0.1 pF
▪ resting membrane potential − 39.6 ± 0.3 mV
Integrin redistributed around the nucleus [32]
Ca2+ inflow (probably through VGCC)
No significant increase in intracellular Ca2+
Functionality has also been regulated with ES in vitro
(Table 2). Both macrophages and fibroblasts are highly
plastic cells that change their phenotype and associated
function as the healing signs of progress [62,68]. Direct
current or low frequency ES triggers Ca2+ intake
(Table 3), and impacts cytokine, enzyme, and matrix
protein secretion in both cell types [35,64,69–73]. At the
same time, macrophage phagocytosis is more efficient
[64], and the contractility of fibroblasts is increased as
they differentiate into myofibroblasts [70,71,74]. Subse­
quently, myofibroblasts could promote the transition to
the macrophage pro-repair phenotype [62,75,76].
However, while most ES devices aim to recreate the
endogenous EF recorded in the wounds of healthy skin,
it is unlikely that cells in chronic wounds will react the
same as their healthy counterparts [13–18]. Both mac­
rophages and fibroblasts undergo profound changes in
patients prone to chronic wounds [13–18], but their ef­
fects on the electrical properties have not been de­
scribed yet. Membrane-bound voltage-sensitive dyes
[77–79], ion-bound fluorescent dyes [32,64,66], ion
channel blockers [80], microfluidic devices [81], and
patch-clamp electrodes [82–84] could be used to quan­
tify those changes in single cells. While current data
show the pro-healing effects of ES on healthy cells, the
stimulus must be optimized to trigger responses in cells
derived from chronic wound-prone patients or model
organisms.
Conclusion
In this short review, the limitations and opportunities in
the characterization of the electrical stimulus for wound-
healing therapies have been outlined. Moreover, it is
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 Reviewing chronic wound bioelectricity principles Verdes et al.
argued that for chronic wounds, especially the full-
thickness ones where the dermal layer is damaged and
inflammatory processes are dysregulated, it is not suffi­
cient to only estimate the stimulus influencing the
wound-edge keratinocytes, and thus only have a control
mechanism for the epithelial layer. Considering this, a
natural step in advancing wound ES treatment protocols
is targeting the cells that orchestrate dermal repair,
namely macrophages and fibroblasts. While both these
cell types were shown to react to an applied EF, suc­
cessful optimization of devices and treatment protocols
still requires interdisciplinary effort in two major bodies
of work. The first one is in modeling the transduction of
the electric stimulus, from the electrodes to the target
cells, both in vitro and in vivo. Crucially here, better
characterization of the dermal EF must be performed in
order to understand the endogenous microenvironment
in which the macrophages and fibroblasts reside. The
second one is in characterizing the ES response of those
cells derived from patients prone to chronic wounds.
Thankfully, a range of tools is already available.
Computational models, enriched with microscopy-en­
abled detailed geometry, can guide ES application in
vitro, help launch hypotheses of ionic flow within the
skin, and help estimate the cellular electrical properties,
as well as their changes because of aging and chronic
diseases. Validation can be sought with ion distribution
experiments in vitro at a single-cell level, or in ex vivo
skin. Eventually, these efforts will inform and facilitate
the design of a new generation of electroceuticals to treat
chronic wounds.
Conflict of interest statement
None declared.
Acknowledgements
This work was supported by the Henry Royce Institute for Advanced
Materials, funded through Engineering and Physical Sciences Research
Council (EPSRC) grants EP/R00661X/1, EP/P025021/1 and EP/P025498/1,
and by the 4-year Wellcome Trust PhD Programme in Quantitative &
Biophysical Biology.
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