Case study
Tools:

Replacing radiolabelingtechniques with the SURFE2R N1

SURFE 2R N1

Dr. Randy Stockbridge
featured by Nanion Technologies
Dr. Randy Stockbridge is Assistant Professor atcontended with vast and unusual chemical threats throughout

the University of Michigan, Ann Arbor, MI, USA.

evolutionary time and retain a catalog of idiosyncratic

She holds a Bachelor of Arts from Princeton

export proteins to deal with hostile elements. Delving deeply

University 2005, and received her Ph. D. frominto bacterial export systems reveals novel physiologies andthe University of North Carolina in 2010.

structural surprises. By understanding bacteria’s intrinsic

After that she went to Brandeis University forvulnerabilities, we can generate new leads on antimicrobials.

postdoctoral work before coming to Michigan.

We have discovered exporters of two different toxic ions:fluoride (F-) and guanidinium (Gdm+). Fluoride channels areinteresting because they protect bacteria against toxic
environmental fluoride ion, and guanidinium ion exporters,
New ways to explore “old“ targets
In 2018 Nanion Technologies ran a contest where
researchers from all over the world had the chance to wina SURFE2R N1 platform for a 6-month period to perform
transporter research in their own laboratories and with theirown scientific scope. Randy Stockbridge and her researchgroup convinced the Nanion team with their application.
Shortly after, a system was installed, the assays optimized,and Dr. Randy Stockbridge and her team started with theirresearch project at once.
NT: Randy, tell us about your research!
RS: My lab focuses on discovering and understanding
membrane transport proteins.
Membrane transport proteins – channels and energy-coupledpumps – are the molecular gatekeepers of the cell. For
microbes afloat in a hostile environment, these proteins importvital nutrients and export dangerous toxins. Bacteria have

Nanion's SURFE2R N1 platform
allows for the investigation of electrogenic
membrane transport proteins with high sensitivity.
So far over 100 targets have been studied
including symporters, exchangers, uniporters, ion
pumps and ion channels.

which have an interesting evolutionary relationship to
multidrug exporters involved in antibiotic resistance. We havefour big questions:•what do they look like?

•how do they work?

•how did they evolve?

•what is their biological role?

“...given our difficulties developing a transport assay, I was
pessimistic that we were going to be able to move beyond
radiolabeled substrates. Contrary to my expectations,electrophysiology with the SURFE2R N1 platform worked rightaway.“

NT: What techniques are you currently using in your scientificcommercially available radiolabeled substrates. Since manywork?

of the substrates aren’t fluorescent, fluorescence-based
methods would not work, and the currents generated by these

RS: Our lab uses a breadth of biochemical andtransporters were not large enough to see with planar lipid

biophysical techniques to study these proteins, including

bilayer electrophysiology.

electrophysiology, membrane protein biochemistry, x-ray
crystallography, and macromolecular NMR, and since 2018

Because of the sensitivity of the SURFE2R N1 where small

also the SURFE R N1 from Nanion Technologies.

electrogenic currents can be measured, it seemed like the2ideal device for us to measure this protein.
NT: What prompted you to enter the competition to win aSURFE2R N1?

NT: What was the project you had in mind and did the SURFE2R
N1 fulfill your expectations?

RS: We had done the initial characterization of our new familyof transporters using radiolabeled substrate uptake, but we

RS: We wanted to characterize the electrophysiologicalneeded a more general assay in order to expand beyond

characteristics of our new family of transporters to find out
properties such as Km and Vmax of the substrate, and ideally

Automation
Automated measurements using
up to 53 solutions.
Up to 150 data points per day (SURFE²R N1).
Up to 10.000 data points per day (SURFE²R 96SE).

InjectorSubstrate

Electric read-out
No labels, direct measurements.
Real-time and immediate responses.

SampleCapacitive read-out
Voltage-control throughionophores.

Robust and reliable
High sensor stability.
Multiple measurements over hours.
High success rates and reproducibility.

Compounds
Substrate and inhibitor assays; for hydrophobic
compounds and metal ions, negative controls
are essential.
High target range
Symporters, exchangers, ATPases, channels.
Sample flexibility
Native membranes or proteoliposomes. Low
sample consumption <0.1 µg protein persensor.
High protein density may be required.

SensorHigh sensitivity
Huge sensor surface.
Targets down to transport rates of
0.1 s−1. Detects even conformational transitions
and binding.

Solid Supported Membrane (SSM) Electrophysiology Measurements on the SURFE2R Technology
The figure shows the SURFE2R technology wih its benefits and limitations explained.

to screen a set of substrates for transporter activity. However,

NT: Are there other techniques you could have used for thisgiven our difficulties in developing a transport assay, I wasproject? If so, what are they and what are the benefits of thepessimistic that we were going to be able to move beyond

SURFE2R N1 over these other techniques?

radiolabeled substrates. Contrary to my expectations,electrophysiology with the SURFE2R N1 platform worked right

RS: We have been using fluorescence and radiolabeled

away. We saw very robust currents in our pilot experiments

uptake experiments but these were unsuitable in this case.

during the initial training session on the instrument. The data we

Now that we’ve started with solid supported membrane (SSM)

collected in these first experiments with the SURFE2R N1 agreedelectrophysiology, this technique has replaced radiolabeledwith what we already knew about the transporters, andsubstrate experiments in my lab, to everyone’s benefit. We’reafter that we were quickly able to able to push the project innot spending as much money on radiolabeled substrates,

exciting, unexpected directions by screening a large family ofthe experiments are much faster to perform, with fewer

structurally related substrates for transport.

regulatory headaches. We were quickly able to push theproject in exciting, unexpected directions by screening a large

NT: How was the support you received from Nanion?

family of structurally related substrates for transport. And mygraduate student vastly prefers doing SSM electrophysiology

RS: The support we got from Nanion Technologies was reallyexperiments, too!

comprehensive; in addition to an in-depth training session at
the start of the SURFE2R N1 demo period, they worked withmy graduate student to analyze and interpret the data anddevelop the proper controls over the 6-month period that wecollected data with the instrument.

Experiments on the SURFE2R N1 in Dr. Randy Stockbridge’s laboratory. Left, Chris Macdonald is recording thebacterial guanidium transporter. Figure shows SSM (solid supported membrane) electrophysiology measurementsof Gdx transporter reactions on the SURFE2R N1. (A) Raw data traces showing activation by application of 1 mM
Gdm+. In (B) and (C) the concentration dependent current resposes and the corresponding Km-plot are shown.

“Now that we’ve started with solid supported membrane (SSM)
electrophysiology, it has replaced radiolabeled substrateexperiments in my lab, to everyone’s benefit. We’re not spending
as much money, the experiments are much faster to perform, withfewer regulatory headaches. And my graduate student vastly
prefers doing SSM electrophysiology experiments, too! “NT: Do you have any further projects in mind that you would

NT: Would you recommend the SURFE2R N1 platform to otherlike to use the SURFE R N1 for? If so, can you tell us a bit more

researchers studying transporters?

2

about them?
RS: The SURFE2R N1 fills a niche that other transport techniques
RS: I also work on a family of microbial fluoride channels. I’vedo not. I would definitively recommend it.

primarily characterized these proteins using planar lipid bilayerelectrophysiology. However, the SURFE2R N1 is so sensitive that
I can monitor currents at much lower substrate concentrations(a few micromolar). We’re interested in probing channel
function at sub-saturating ion concentrations using the SURFE2R
N1.
I’ve also had several inquiries from colleagues interested
in using the SURFE2R N1 for various applications, including
measuring transporter or ion channel currents in organelles ormembrane vesicles.

Contact InformationDr. Randy Stockbridge
Assistant Professor
Department of Biophysics & Department of Molecular,
Cellular, and Developmental Biology
University of Michigan
930 N. University Ave.
Ann Arbor, MI 48109-1055
stockbr@umich.edu
https://sites.lsa.umich.edu/randystockbridge/

SURFE2R Team at Nanion
Dr. Maria Barthmes
Ganghofer 70a80339 Munich
Germany
Email: maria.barthmes@nanion.de
Phone: +49 89 219095 068
Web: www.nanion.de

Acknowledgments
We thank Dr. Stockbridge for letting us use images anddata, and for the very nice conversation!

Nanion Technologies GmbH I Ganghoferstraße 70a, 80339 Munich, Germany+49 89 2190 95-0 I info@nanion.de I www.nanion.de