Quantum Super-resolution Imaging in Fluorescence Microscopy Osip Schwartz, Dan Oron, Jonathan M. Levitt, Ron Tenne, Stella Itzhakov and Dan Oron Dept. of Physics of Complex Systems Weizmann Institute of Science, Israel FRISNO 12, Ein Gedi (February 2013) Microscopy and resolution Resolution of far-field optical microscopes is limited by about half wavelength. (Ernst Abbe, 1873) Workarounds: • Nonlinear optical methods: use nonlinear optical response to produce narrower point spread function • Stochastic methods: use fluorophores turning on and off randomly • Quantum optics? o Multi-photon interference o Entangled images o Sub shot noise imaging o Resolution enhancement? Slide 2 of 18 Afek et al., Science 328 (2010) Walther et al., Nature 429 (2004) Boyer et al., Science 321 (2008) Brida et al., Nat. Photonics 4 (2010) Quantum super-resolution • Quantum Limits on Optical Resolution • Wolf equations for two-photon light • Quantum Imaging beyond the Diffraction Limit by Optical Centroid Measurements • Quantum spatial superresolution by optical centroid measurements • Quantum imaging with incoherent photons, Thiel et al., PRL 2007 • Sub-Rayleigh quantum imaging using single-photon sources Thiel et al., PRA 2009 • Sub-Rayleigh-diffraction-bound quantum imaging, • Sub-Rayleigh Imaging via N-Photon Detection, Quantum light Slide 3 of 18 Kolobov, Fabre, PRL2000 Saleh et al., PRL 2005 Object M.Tsang PRL 2009 Shin et al., PRL 2011 Giovannetti, PRA 2009 Guerrieri et al., PRL 2010 Imaging system Light detector Quantum emitters Classical light Quantum light Quantum emitters Imaging system Light detector What if we had an emitter that would always emit photon pairs? S.W. Hell et al., Bioimaging (1995) Slide 4 of 18 Multi-photon detection microscopy Photon pair τ1 τ2 τ1>>τ2 cascaded emitters Imaging system Photon pair detector Point spread function: h2phot(x) = h2(x) Spatial distribution of photon pairs carries high spatial frequency information (up to double resolution) Similarly, in N-photon detection microscopy hNphot(x) = hN(x) Slide 5 of 18 Antibunching microscopy Observations of antibunching: Number of photons emitted after excitation: Organic dyes: W. Ambrose et al. (1997) Quantum dots: B. Lounis et al. (2000). 1 NV centers: R. Brouri et al. (2000). 0.5 Classical 0 Fluorescence intensity autocorrelation g(2) 0 1 2 Pair source 3 1 0.5 0 0 1 2 3 Single photon source 1 0.5 0 10 μs interval between pulses Slide 6 of 18 0 1 2 3 Instead of actual photon pairs, consider ‘missing’ pairs. Antibunching-induced correlations Two adjacent detectors in the image plane: x0 For individual fluorophore: For multiple fluorophores: Sum over fluorophores Slide 7 of 18 Emitters Fluorescence saturation CdSe / ZnSe / ZnS quantum dots 1 0.8 0.6 0.4 0.2 0 450 Slide 8 of 18 500 550 600 Wavelength, nm 650 Schwartz et al.,ACS Nano 6 (2012) At 1 kHz: Slide 9 of 18 Schwartz et al.,ACS Nano 6 (2012) Photon counting with a CCD Pixel signal distribution Read noise threshold CCD ADC units More signal Slide 10 of 18 Less noise arXiv:1212.6003 Computing correlations 2nd order: Quantifies the missing pairs 3rd order: • compute correlations for all pixel configurations • Fourier-interpolate the resulting images • Sum the interpolated images Slide 11 of 18 Missing 3-photon events (except those due to missing pairs, already accounted for) arXiv:1212.6003 Antibunching with a CCD Third order: g(3)(τ1, τ2)= =<n(t)n(t+τ1)n(t+ τ2)> τ, ms τ, ms τ2, ms g(2)(τ)=<n(t)n(t+ τ)> Classical signal τ2, ms Second order autocorrelation function: Quantum dot τ1, ms Slide 12 of 18 τ1, ms arXiv:1212.6003 1 8000 2 Fluorescence image 6000 3 4 4000 5 2000 6 7 0 2 4 6 8 10 12 1 500 2 400 3 300 4 5 200 6 100 7 0 2 4 6 8 10 12 1 25 2 20 3 15 4 10 5 Slide 13 of 18 6 5 7 0 2 4 6 8 10 12 arXiv:1212.6003 1 8000 2 Fluorescence image 6000 3 4 4000 5 2000 6 7 0 2 4 6 8 10 12 1 2nd order antibunching 500 2 400 3 300 4 5 200 6 100 7 0 2 4 6 8 10 12 1 25 2 20 3 15 4 10 5 Slide 14 of 18 6 5 7 0 2 4 6 8 10 12 arXiv:1212.6003 1 8000 2 Fluorescence image 6000 3 4 4000 5 2000 6 7 0 2 4 6 8 10 12 1 2nd order antibunching 500 2 400 3 300 4 5 200 6 100 7 0 2 4 6 8 10 order antibunching 25 2 20 3 15 4 181 nm FWHM (x1.50) 10 5 Slide 15 of 18 216 nm FWHM (x1.26) 12 1 3rd Resolution: 271 nm FWHM 6 5 7 0 2 4 6 8 10 12 arXiv:1212.6003 Optical sectioning Defocused image of a quantum dot: Fluorescence imaging 400 300 200 2nd order antibunching imaging 400 300 200 15 10 5 15 10 5 1 1 0.8 0.8 0.6 Optical signal integrated over the field of view: 0.4 0.6 0.4 0.2 0.2 0 Slide 16 of 18 -1 0 -0.5 0 -1 0.5 -0.5 1 0 0.5 Defocusing, μm 1 Summary •Far-field super-resolution imaging demonstrated by using quantum properties of light naturally present in fluorescence microscopy •The experiment was performed with commercially available equipment, at room temperature, with commonly used quantum dot fluorophores •With further development of detector technology, antibunching imaging may become feasible as a practical imaging method Slide 17 of 18 The team Jonathan M. Levitt Stella Itzhakov Ron Tenne Slide 18 of 18 Zvicka Deutsch Dan Oron A D 8000 1 1 2 8000 2 6000 3 6000 3 4 4 4000 5 4000 5 2000 6 7 7 0 2 4 6 8 10 12 B 0 2 500 1 2000 6 4 6 8 10 12 E 1 2 400 2 3 300 3 500 400 4 300 4 200 5 100 6 0 7 2 4 6 8 10 5 200 6 100 7 0 12 2 4 6 8 10 12 F C 1 25 2 1 25 2 20 3 20 3 15 4 10 5 15 4 10 5 6 5 6 5 7 0 7 0 2 Slide 19 of 18 4 6 8 10 12 2 4 6 8 10 12 Superresolved images Reconstructed high resolution images Regular (photon counting) image Slide 20 of 18 Second order correlations Third order correlations Superresolved images Slide 21 of 18 arXiv:1212.6003 Superresolved images Slide 22 of 18 Superresolved images Slide 23 of 18 A 4 6 8 277 267 262 286 284 269 258 244 258 272 280 273 274 276 274 271 288 268 286284 262 253 256 249271295 259 261 262 266 12 276 10 B 304 15 4 266 224203 206 207 219 213 213 6 215 216 217 6 2 212 213 227 0 200 205 212 210 210 4 6 180 4 186 5 180 162 191 181 183 194 171 175 4 210 10 175 181 157 166 175 0 150 175 188 185 185 2 181 5 178 188 189 180 189 7 Slide 24 of 18 200 180 180 186 168 189 181 F 15 185 183 171 171 186 6 230 183 186 171 196 225 0.18143 181 196 178 191 189 205 188 186 178 176 185168 188 175 189 171 178 186 183 194 186 178 220 12 181 171 185 178 215 166 181 181 173 188 181 185 10 210 PSF width (nm) 223 8 191 185 3 310 E 217 2 300 216 220 7 186 290 215 217 1 280 8 217 221 C 270 4 220 2 260 0.21557 212 217 217 217 250 25 215 5 2 0 240 224 215 223 219 4 6 PSF width (nm) 20 206 215 217 219216 221 8 209 215 215 10 271 219 212 3 0.27182 268 269 260 263 215 2 12 270 262 268 1 213 262 267 270 267 282 278 291 263 14 259 266 253 274 287 273 266 299 274 276 283 251 5 277 286 282 282 271 279 253 268 277 264 255 271 279 283 D 273 272 273 283 260 253 10 270 283 268 259 256 279 292 286 266 280 273 262 277 289 267 283 258 2 267283 292281 282 6 8 10 12 160 170 180 190 PSF width (nm) Quantum super-resolution Conceptual difficulty: an absorptive grating with subwavelength period acts as an attenuator for every photon • Transmitted light contains no information on the grating phase or period • Any linear absorber mask is a superposition of gratings • High spatial frequency components of the mask are lost Slide 25 of 18