Segmented Field OFFGEL® Electrophoresis
---- Supporting Information ----
E. Tobolkina1, F. Cortés-Salazar1, D. Momotenko1, Julien Maillard2, Hubert H.Girault1
1
Laboratoire d’Electrochimie Physique et Analytique, Ecole Polytechnique Fédérale
de Lausanne (EPFL), Station 6, CH-1015 Lausanne, Switzerland
2
Laboratory for Environmental Biotechnology, Ecole Polytechnique Fédérale de
Lausanne (EPFL), Station 6, CH-1015 Lausanne, Switzerland
*
CORRESPONDING AUTHOR FOOTNOTE
E-mail:
hubert.girault@epfl.ch
Telephone number:
+41-21-693 3145
Fax number:
+41-21-693 3667
SI- 1: Finite element simulations
In order to reduce computational efforts the solution was computed sequentially, i.e.
solving mass-transport equations on top of the stored solution containing electric field
distribution. The mesh size was adjusted down to the value of 10 m at the corners of
the wells and electrode edges.
Figure SI-1. The mesh mapped over computational domain used in FEM modeling.
SI-2: Numerical simulation of protein migration and electric field distribution in
the OFFGEL device
The constants used in finite element simulations for numerical analysis are
summarized in the table below.
Table 1. Constant parameters for the numerical simulations.
Constant
Value
Units
R (gas constant)
8.31
J/mol*K
T (temperature)
298
K
96485
C/mol
5.5*10-6
S/m
9*10-9
m2/s
OH-(diffusion coefficient of OH-)
5.27*10-9
m2/s
D (diffusion coefficient of α-lactalbumin)
1.06*10-10
m2/s
F (Faraday constant)
δDI (water conductivity)
H+ (diffusion coefficient of H+)
pKa values for amino acids were from the Handbook of Chemistry and Physics
(http://www.hbcpnetbase.com/)
Due to numerical instabilities for the convergence of Nernst-Planck equation the
overall potential difference was set to 1 V. The choice of the potential difference is
arbitrary for comparison of the efficiency and separation rate of different
electrophoretic setups and voltage programs as the only requirement is to maintain
equal conditions for electrophoretic separations.
Table 2. The potential programs used in numerical simulations.
Electrode No.
1
2
3
4
5
6
7
Multielectrode
setup
Optimized
potential
program for
general protein
mixtures (V)
0
0,211
0,359
0,453
0,606
0,770
1
Multielectrode
setup
Adjusted
voltage
program
for
separation
of
proteins with
acidic pI (V)
0
0,05
0,125
0,225
0,35
0,5
0,675
Twoelectrode
setup
Potential
program used
in twoelectrode setup
(V)
0
-
-
-
-
-
0,675
SI-3: OFFGEL electrophoresis with a multi-electrode setup
All the fractions taken from each well after OFFGEL separation employed for 3 hours
using multi-electrode setup were analyzed with MALDI-mass spectrometry. As it
seen from the Figure SI-3, no proteins were observed in the wells No.2, 4 and 6
showing the efficient separation made by proposed approach.
Figure SI-3. Mass spectra of fractions taken after OFFGEL electrophoresis
performed using multi-electrode setup, from each of the seven wells and deposited on
a steel target plate with sinapinic acid (SA) matrix.
SI-4: OFFGEL electrophoresis with a multi-electrode setup
After the OFFGEL separation performed with two- and multi-electrode setups using
potential programs, summarized in Table 1, the gel strips from all of four experiments
were stained by standard protocol using Coomassie blue solution.
A
B
C
D
Figure SI-4. Coomassie blue stained gel strips after OFFGEL electrophoresis
performed with A) Multi-electrode setup B) Two-electrode setup (using multielectrode device) C) Two-electrode setup (using Agilent Fractionator 3100 and an
optimized potential program) D) Two-electrode setup using standard method
recommended by Agilent Technologies in their operator manual.