The 5th Korea-U.S. Nano Forum
Eletrokinetic Protein Preconcentration
Using Nanochannels Formed By Weak
Bonding of PDMS Membrane with Glass
Substrate
Sun Min Kim
Dept. of Mechanical Engineering,
Inha University
2008. 4. 18.
INHA University
Department of Mechanical Engineering
University of Michigan, Ann Arbor
Department of Mechanical Engineering
Lab on a chip (Laboratory on a chip)
Lab on a Chip
Sample acquisition
Sample preparation
Analyte Concentration
Analyte detection
Analyte Amplification
Why concentrate samples?
– Sensitivity: Concentration in sample often less than detection limits of
instruments
– Waste: Most samples are > 1 mL in size, but only nanoliters can be analyzed
at a time on a microchip
Bio-Micro Fluidics Lab.
INHA University
Department of Mechanical
Engineering
-2-
Simple Glass/PDMS Preconcentrator
GND
PBS Buffer
1
M
2
M’
4
20µm
–
PDMS
Separation channel
Gap:20μm
3
M
PBS + Protein sample
M’
+
Glass
5
PDMS
Glass
+ HV
• Channel dimensions
Chevron channels (W 40µm× D 18µm, L=16mm),
Separation channel (W 30µm× D 18µm, L=40mm)
• Microchannels are created by casting PDMS over a mold
(SU-8 on a Si substrate).
Bio-Micro Fluidics Lab.
INHA University
Department of Mechanical
Engineering
-3-
Experimental Results
GND
PBS Buffer
1
2
Separation channel
Gap:20μm
3
100µm
5
4
PBS + Protein sample
PDMS
Glass
+ HV
Experimental conditions
•Protein sample : Fluorescein isothiocyanate (FITC) conjugate bovine
serum albumin (BSA) and ovalbumin (OVA)
•Buffer : Phosphate-buffered saline buffer (10mM, pH 7.4) and
Phosphate buffer (20mM, pH 7.2)
•O2 plasma treatment of the PDMS
(100W, 200mTorr, 3min)
•Applied electric potential: 100, 200, 300V
Bio-Micro Fluidics Lab.
INHA University
Department of Mechanical
Engineering
-4-
Experimental Results (cont’d)
Concentration of FITC-BSA in the preconcentration zone
3000
1pM
100pM
10nM
5µM standard
10µM standard
20µM standard
Fluorescence Intensity (A.U.)
2500
2000
1500
1000
500
Preconcentration Zone
0
0
5
10
15
20
25
30
35
Time (min)
Concentration achieved up to 106 –fold
Bio-Micro Fluidics Lab.
INHA University
Department of Mechanical
Engineering
-5-
Separation of preconcentrated proteins
Bio-Micro Fluidics Lab.
INHA University
Department of Mechanical
Engineering
-6-
Physical Mechanism: Hypothesis and Verifications
“nanoscale channels” formed between the PDMS and the glass due to the
weak, reversible bonding
1. No preconcentration occurs in PDMS/PDMS and irreversibly bonded PDMS/glass
(stronger bond) chips.
2. Nanochannels work as a cationic selective membrane due to the Ion exclusionenrichment effect (EEE) caused by electrical double layer (EDL) overlapping.
3. Electroosmotic flow (EOF) dominates over electrophoresis (EP)
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–
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–
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–
– –
– –
INHA University
Department of Mechanical
Engineering
-7-
–
–
–
–
– –
– –
–
Bio-Micro Fluidics Lab.
Ⓘ
Ⓘ Ⓘ
4
λD
Ⓘ
⊕⊕⊕⊕⊕⊕⊕ Hn
⊕⊕⊕⊕⊕⊕⊕⊕ ⊕⊕ ⊕ ⊕ ⊕ ⊕ ⊕⊕⊕⊕⊕⊕⊕⊕
Glass
Ⓘ
M’
Ⓘ Ⓘ
Ⓘ
3
PDMS
Ⓘ ⒾⒾ
Ⓘ
ⒾⒾ
5
⊕⊕⊕⊕⊕⊕⊕⊕
⊕
⊕
Ⓘ ⒾⒾ
Ⓘ Ⓘ
⊕⊕⊕⊕⊕⊕⊕⊕
⊕
⊕
– Hm
Ⓘ Ⓘ Ⓘ
2
–
M
Ⓘ
1
M’
–
M
EP
–
EOF
+
Experimental Verifications
Verification of the cationic selectivity of the nanochannel
– Sample: 1µM Rhodamine 123 dye (cationic) in PBS buffer (bottom)
Bio-Micro Fluidics Lab.
INHA University
Department of Mechanical
Engineering
-8-
Experimental Verifications (cont’d)
Verification of the dominant electroosmotic flow (EOF)
– Sample: 1µM FITC-BSA in PBS buffer (Top)
Bio-Micro Fluidics Lab.
INHA University
Department of Mechanical
Engineering
-9-
Conclusions
• PDMS / Glass chip for protein preconcentration
was designed and fabricated.
• Preconcentration of labeled BSA (FITC BSA) has
been achieved up to 106 – fold.
• Preconcentrated protein was injected and
separated in a separation column.
• “Nanoscale channels” formed between the PDMS
and the glass due to the weak, reversible bonding
works as a cationic selectivity membrane by Ion
exclusion-enrichment effect (EEE).
Bio-Micro Fluidics Lab.
INHA University
Department of Mechanical
Engineering
-10-