Innovative sensors and systems for the molecular Point-of-Care diagnostics

Transcription

1 Innovative sensors and systems for the molecular Point-of-Care diagnostics AIT Austrian Institute of Technology GmbH Health & Environment Department Molecular Diagnostics Contact

2 AIT Austrian Institute of Technology GmbH AIT headquarter Schönbrunn palace Largest research center in Austria (~1150 employees) Half public, half private Five departments: Energy Mobility Safety & Security Health & Environment Innovation Systems 2

3 Organizational chart of AIT

4 Business Unit Molecular Diagnostics Biomarker Development Assay Development Diagnostic Biosensors Molecular Diagnostics Systems Integration Point-of-Care Bioinformatics 4

5 Automated immunoassays for monitoring at intensive care units Project leader Dr. Johannes Peham PhD topic of Helene Zirath 5

6 Automated immunoassays: microfluidic design Disposable thermoplastic chip Reaction chamber: 20 µl, air-free filling, storing of magnetic beads Chip-integrated micro-syringes: Actuation of detection antibody and sample, onchip storage of detection antibody Outlet Inlets and outlet for reagents Mixing channel for substrate Reaction chamber (20 µl) with magnetic beads 6

7 Automated immunoassays: analysis process 1. Chip prefilled with magnetic beads, detection antibody and plasma 2. Sample incubation 3 min + wash 3. Detection antibody-biotin 3 min + wash 4. SA poly-hrp 3 min + wash 5. Chemiluminescent substrate (mixed on-chip) & read signal Zirath H, Proc MicroTAS, 2014 Zirath H, Proc Eng,

8 Luminescence Units Automated immunoassays: measurement range On-chip IL-8 ELISA IL-8 biomarker in 10 % plasma (pg/ml) On-chip IL-8 ELISA Signal blank Limit of detection LOD on-chip: 10 pg/ml IL-8 detected at concentrations between 10 pg/ml and 2000 pg/ml within only 30 minutes These concentrations correspond to the clinical need by requiring only 3μL blood plasma for the measurement Zirath H, Proc MicroTAS, 2014 Zirath H, Proc Eng,

9 Summary Automated immunoassays for monitoring at intensive care units Rapid <30 minutes Compact demonstrator: 20x20cm testchip: 2x2cm Outlook: Validate with patient samples Develop multiplex system Sensitive <= 50µL whole blood LOD ~ 10pg/mL Cost-effective reagents/marker 1.1 cartridge ~<1 device < 1000 planned Patent pending EP Project leader Dr. Johannes Peham johannes.peham@ait.ac.at PhD topic of Helene Zirath 9

10 Nanoparticle-based homogeneous biosensing Project leader Dr. Jörg Schotter 10

11 Nanoparticle-based homogeneous biosensing Mix & measure detection principle sample + functionalized nanoparticles cuvette 11

12 Nanoparticle-based homogeneous biosensing Mix & measure detection principle Analyte molecule specifically binds to functionalized nanoparticle 12

13 Nanoparticle-based homogeneous biosensing Mix & measure detection principle Analyte molecule specifically binds to functionalized nanoparticle On analyte binding, a measureable physical property of the nanoparticle changes 13

14 Nanoparticle-based homogeneous biosensing Optical detection of nanoparticle phase lag in external rotating magnetic field hybrid magnetic-core / Au-shell nanorods absorption allows optical detection of nanoparticle orientation which depends on surface coverage by selectively bound target molecules Illustration by Darragh Crotty, S. Schrittwieser et al., ACS Nano 6 (2012) 791

15 Nanoparticle-based homogeneous biosensing Measurement setup & typical raw signals PC sample audio amplifier Lock-in amplifier photodetector small coil shunt signal (V) voltage (V) photodetector signal (V) 2.34 small coil shunt voltage (V) 2 laser diode photodetector coil pairs Laboratory setup φ time (ms) 2f-optical signal due to symmetry of nanorods Phase lag is measured by lock-in amplifier and re-calculated to 1f -1 15

16 Summary Nanoparticle-based homogeneous biosensing Simple mix & measure technique Minimal sample preparation Method applicable sensing in serum and saliva Fast Reduced incubation time due to 3d diffusion Continuous monitoring of binding events (Real-time measurements) Cost-effective Easy to integrate & simple instrumentation Small sample volumes Ideally suited for point-of-care applications Patent status Austrian AT European EP Chinese CN US application US Other applications Results can be modeled and understood by theory Size analysis of bound proteins Measurement of kynetic parameters possible S. Schrittwieser et al., Small 10 (2014) 407 Project leader Dr. Jörg Schotter 16

17 Integrated optical waveguide biosensing Project leader Dr. Rainer Hainberger 17

18 The photonic sensing principle: MZI waveguide side view Bio-sensitive layer on top of waveguide provides selective binding and enrichment of biomolecules on sensor surface Light intensity analyte sensing layer n sl core layer n wg binding events change phase velocity of light in sensing arm Intensity Concentration substrate n s waveguide front view w~600nm Si 3 N 4 H~250nm SiO 2 18

19 The photonic sensing principle: MZI 19

20 light power (µw) The photonic sensing principle: MZI 2,0 1,5 1,0 interference 0,5 0,0 1,00 1,25 1,50 1,75 2,00 2,25 time (min) ~1cm bound target molecules biofunctional layer with capture molecules 20

21 Measurement setup 21

22 Miniaturization: optoelectronic integration =850nm input grating optical sensor element output grating silicon dioxid µm Si-photodiodes optoelectronic layer Si 3 N 4 waveguides fabricated by CMOS compatible process TSV silicon substrate 22

23 Miniaturization: optoelectronic integration SOI PECVD-Si 3 N 4 Silicon as waveguide material Silicon nitride (Si 3 N 4 ) as waveguide material Wavelength of 1310nm Wavelength of 850nm Requires special SiGe photodiodes Co-integration with optoelectronics requires wafer-bonding process Integration of standard Si photodiodes possible Direct PECVD deposition on Si-optoelectronics possible 23

24 optical power (dbm) Experimental results: low-loss 1cm 0.86dB/cm (<18% of optical power lost over 1 cm) =850nm;TM 24

25 normalized power (1) Experimental results: sensitivity & biosensing Si 3 N 4 wire waveguides thickness 250nm 1cm 1,0 0,9 0,8 0,7 0,6 0,5 0,4 0,3 0,2 0,1 0,0 with NaCl-solution n=~9e time (sec) S-Protein/S-Peptide measurement

26 Compact PECVD-Si3N4 MZI spirals (L=3cm) 470µm

27 Technology for OCT applications Austrian national project aiming to realize a prototype with following characteristics: Scalable Suitable for mass production Using a light source with a wavelength of 850nm Integrated detector on chip Integrated analog optoelectronics High efficient coupling and waveguiding

28 Summary Integrated optical waveguide biosensing Mach-Zehnder Interferometer biosensing Suitable for proteins, peptides, DNA, antibodies CMOS compatible low-loss Si 3 N 4 waveguide platform Suitable for point-of-care applications Other possible applications: OCT, Telecommunications Reduction of sensing area by integrating spiral-shaped waveguides Multiplex sensing Project leader Dr. Rainer Hainberger rainer.hainberger@ait.ac.at 28

29 Micro- & nanotechnology equipment Microfluidic milling machine LPKF Protomat Wire bonder F+K Delvotec, manual operation Evaporation chamber Leybold Univex 450, 2 thermal sources, 1 e-gun source Argon ion beam etching Roth+Rau, IonSys 500, 4 ion source Hiden mass spectrometer for end point detection Plasma asher Diener Electronics, Femto Optical lithography EVG 620 mask aligner, 4 SEM & e-beam lithography Zeiss Supra 40, Raith e-beam writing tool Cleanroom 42m², resist processing and wet chemical etching Atomic Force Microscope Molecular Imaging, Pico Plus Profilometer Tencor, Alpha Step Magnetron sputtering Leybold, Univex 450 C cluster system, 11 Targets, 4, DC/RF sputtering Materials inkjet printer FujiFilm Dimatix DMP-2831

30 Electrical, magnetic & optical characterization Analytical Probe System SUSS PM5, electrical DC-HF measurements on 6 wafers or single substrates, compatible with two-axis (in-plane) magnetic field application for magnetoresistance characterization Precision Semiconductor Parameter Analyzer Agilent 4156C, for frequency-dependent LRC-measurements Probe station for magnetoresistance characterization in arbitrary magnetic field direction (up to 40 mt) Setup for linear AC and rotating magnetic fields up to 2 khz frequency and 5 mt amplitude including optical detection Setup for optical waveguide characterization various laser sources (850nm, tunable lasers nm, nm), polarization control, automated alignment & automated liquid handling Monochromomator VIS, NIR, IR SUSS PM5 Analytical Probe System

31 Photonic design & characterization Photonic simulation cluster: 256 cores with 488GB RAM 3D FDTD simulation of photonic wire Bragg reflector 1TB high performance SSD array and 5TB raid-5 HDD storage with automated backup 2D/3D finite difference time domain simulation packages variational mode matching tools for eigenmode calculation Floquet-Bloch solver for simulating waveguide grating couplers Fully equipped set-up for waveguide device characterization and biosensing experiments Temperature controlled sample holder and programmable fluid control system with five fluid reservoirs 31

32 Thank you for your attention!!! Contact AIT Austrian Institute of Technology Health & Environment Department Molecular Diagnostics