The bovine serum albumin (BSA)-polystyrene (PS) interface layer is laser photo-activated at 157 nm for site-selective multiple target–protein immobilization. The 5-15 nm photon-induced interface layer has different chemical, wetting, and stiffness properties than the PS photon processed surface. The irradiated areas exhibit target-protein binding, followed by localized probe-target protein detection.
The photon-induced chemical modification of the BSA-PS interface layer is identified by:
(1) Morphological, imaging and analysis of surface parameters with Atomic Force Microscopy (AFM).
(2) Spectroscopic shift (4 cm-1), of the amide I group and formation of new C=N, NH2, C-O, C=O, O-C=O groups following irradiation, identified with Attenuated Total Reflection Fourier Transform Infrared (ATR-FTIR) spectroscopy.
(3) The different hydrophilic/hydrophobic and force-distance responses of the bare PS and BSA-PS surfaces.
Near field edge diffraction (Fresnel) fluorescence imaging specifies the threshold photon energy and the fluence required to detect optically the protein detection on the photon-induced BSA-PS interface layer. By approximating the Fresnel integrals with analytical functions, the threshold photon energy and the fluence are expressed as the sum of zero, first and second-order harmonic terms of two characteristic diffracted modes and they are specified to be 8.73×10-9 J and 623 Jm-2 respectively. Furthermore, a bioarray of three probe-target proteins is fabricated with 1.5 micron spatial resolution using a 157 nm laser micro stepper. The methodology eliminates the use of intermediate polymer layers between the blocking BSA protein and the PS substrate in bioarray fabrication.
Left: ATR-FTIR spectrum of non-irradiated and irradiated with 20 laser pulses bare PS substrate from 539 to 5000 cm-1.
RIght: ATR-FTIR spectrum of the aromatic C-H stretching vibration mode at 3082.1 cm-1 of the PS substrate irradiated with 1 (1p), 10(10p), and 20(20p) laser pulses respectively.
Left: ATR-FTIR spectrum of the 1701.6 cm-1 band of bare PS with 0(0p), 1(1p), 10(10p), and 20(20p) laser pulses, indicating activation of the C=O group by atmospheric oxygen after irradiation. The spectra are normalized using the spectral range from 1490 to 1630 cm-1.
Right: ATR-FTIR spectrum of the bare PS substrate from 1500-2300 cm-1irradiated with 1(1p),10(10p) and 20(20p) laser pulses respectively. The peaks at 1543, 2101.1, 2336.2, and 2361 cm-1 correspond to the presence of NH2 from the scission of [-N=C=N-] and C-N stressing modes respectively.
Left:ATR-FTIR spectrum of the 1635-1700 cm-1 band of BSA -PS following irradiation with 0 (BSA-PS non exposed), 1(1p), 2(2p), 10(10p), 20(20p) laser pulses.
Right: ATR-FTIR spectrum of the 3200-3370 cm-1 band of BSA-PS with a similar response to the band’s peak at 3300 cm-1.
Left: Intensity distribution of the fluorescence image taken with a CCD camera across one micro-spot, from one to thirty laser pulses.
Right: Intensity distribution of the fluorescence image taken with a CCD camera across one micro-spot, fabricated with 40 to 1080 different laser pulses.
Relevant Publications
Protein immobilization and detection on laser processed polystyrene surfaces.
E. Sarantopoulou, P. S. Petrou, Z. Kollia, D. Palles, N. Spyropoulos-Antonakakis,
S. Kakabakos and A. C. Cefalas,
J. Appl. Phys. 110(6), 064309 (2011).
DOI:10.1063/1.3627160
157nm laser ablation of polymeric layers for fabrication of biomolecule microarrays.
A.M. Douvas, P.S. Petrou, S.E. Kakabakos, K. Misiakos, P. Argitis, E. Sarantopoulou,
Z. Kollia and A.C. Cefalas
Anal. Bioanal. Chem., 381 (5), 1027 (2005).
DOI:10.1007/s00216-004-2985-3
Conference Presentations
Nano-engineering of bio-arrays at 157 nm.
A.C. Cefalas*, E. Sarantopoulou, Z. Kollia, P. Petrou and S. Kakabakos,
11th Trends in Nanotechnology International Conference, TNT 2010, Braga,
Portugal, 06-10 September, 2010.
Nano-engineering of BIO-ARRAYS with Vacuum Ultraviolet Light.
E. Sarantopoulou, Z. Kollia, A. C. Cefalas, A. M. Douvas, M. Chatzichristidi, P. Argitis,
K. Misiakos, Z. Petrou and S. Kakabakos,
EMRS 2007, Strasburg France, May 28 – June 1, (2007).