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Adaptive optical biocompact disk for molecular recognition Leilei Peng, Manoj M. Varma, Fred E. Regnier, David D. Nolte Applied Physics Letters, May 2005 Purdue University Journal Club – 11-03-05 Emre Özkumur Electrical and Computer Engineering Department Boston University outline Background for phase sensitive interferometric detection Details of how the method works Phase modulation with a BioCD Conversion of phase modulation to amplitude modulation Fabrication of the BioCD Results Sensitivity Background for bio-detection; specific, non-specific binding Molecular data Phase sensitive interferometry ε1 E1e ε2 E2 e j 2 jn d 2 d Measure d(n-1) d I E1e 2 j d E2 e 2 2 jn d 2d I 0 I cos (n 1) Phase sensitive interferometry_2 I Most sensitive region: quadrature Satisfied when d(n-1)=λ/4 or… 0 π/2 Ø 2π by having a timeindependent π/2 phase difference between reference and signal beams Phase modulation with a BioCD • 4” diameter • 1024 wedgedspoke pattern of biomolecules • spun at 50Hz, pattern is transferred to probe beam as phase modulation at 51.2kHz Phase modulation -> Amplitude modulation Phase modulated probe and reference beams are sent into a Photorefractive Quantum Well device PRQW E 2’ E 1” E1 E2 E 1’ E 2” - Fringes change the effective refractive index of the material - Because of the periodic change in “n”, device acts as a diffraction grating Why use PRQW? 2 E1 ( E2 , L) e i 0 E1 (0) ( ) E2 (0) exp i P ( ) d (t )( n 1) 2 Photorefractive phase shift Excitonic spectral shift - Quadrature condition is satisfied by adjusting the excitonic spectral shift by changing the applied voltage to PRQW and wavelength of the beam P ( ) - Grating can adapt to slowly varying phases, but not to rapid changes => low frequency variations (environmental & mechanical disturbances) of the phase compansated, high frequency pass without changing - Resultant intensity amplitude modulated with 51.2kHz; total intensity is measured and sent to a spectrum analyzer or lock-in amplifier Fabrication of the BioCD - Ink gel stamping method Sensitivity of the system To test the system sensitivity, a phase modulator was used instead of BioCD Phase modulator was used to calibrate the data, to extract the height information Sensitivity of the system Phase modulator created Øpp=2.1*10-2 rad, corresponding to: pp d (n 1) 2.8nm 2 (optical thickness for λ=830nm) Patterned Bovine serum albumin (BSA) optical thickness measured to be: d(n-1)=0.6nm for which AFM measured 2.5±0.7nm for the height difference, confirming the refractive index of BSA to be n=1.2±0.1nm Signal-to-Noise NEOT: noise equivalent optical thickness NL: Noise contribution of the local oscillator NP: Phase noise from the disk S d d 2 2 N N L N P NEOT 2 2 At 51 kHz modulation frequency NP dominates, NEOT=0.3nm Higher sensitivity can be achieved by narrowing the detection bandwidth (3kHz) Background for bio-detection data BioCD was implemented for an antigen-antibody molecular recognition experiment For simplicity: Label Label Label Label “mouse IgG antigen” = M “anti-mouse IgG antibody” = m “rabbit IgG antigen” = R “anti-rabbit IgG antibody” = r System should be able to detect the bindings of interest (specific bindings) which are M-m, R-r, and should ignore non-specific bindings such as M-r or R-m Preperation of the samples on CD First, M-antigen 1024spoke pattern was printed on the disk Free surfaces (unprinted areas) were covered with BSA Then the disk was divided 4 annular tracks (A-D) Molecular data_1 Non-specific binding was tested by incubating track C with r-antibody Specific binding was tested by incubating both tracks B and C with m-antibody Molecular data_2 This time, after M-antigen 1024 spoke pattern was printed on the cd, R-antigen was used as free-surface saturating reagent instead of BSA Tracks B and C exposed to R-antibody, whereas tracks C and D exposed to M-antibody When no antibody agents present, the homodyne signal is the same as noise When both antibodies bind in track C, homodyne signal diminishes, meaning that the heights cancel each other Conclusion First adaptive interferometric BioCD for molecular recognition was demonstrated Sensitivity = 0.3nm Higher sensitivity can be achieved by Narrowing the detection bandwidth Shifting to a reflection geometry that will not be sensitive to density fluctuations in the glass