Abstract

Phase imaging with a high-resolution wavefront sensor is considered. This is based on a quadriwave lateral shearing interferometer mounted on a non-modified transmission white-light microscope. The measurement technology is explained both in the scope of wave optics and geometrical optics in order to discuss its implementation on a conventional microscope. In particular we consider the effect of a non spatially coherent source on the phase-image signal-to-noise ratio. Precise measurements of the phase-shift introduced by microscopic beads or giant unilamellar vesicles validate the principle and show the accuracy of the methods. Diffraction limited images of living COS-7 cells are then presented, with a particular focus on the membrane and organelle dynamics.

© 2009 Optical Society of America

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  6. M. R. Arnison, K. G. Larkin, C. J. R. Sheppard, N. I. Smith, and C. J. Cogswell, "Linear phase imaging using differential interference contrast microscopy," J. Microscopy 214(1), 7-12 (2004).
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  17. C. Curl, C. J. Bellair, P. J. Harris, B. E. Allman, A. Roberts, K. A. Nugent and L. M. D. Delbridge, "Single cell volume measurement by Quantitative Phase Microscopy (QPM): A case study of Erythrocyte Morphology," Cellular Physiology Biochem. 17, 193-200 (2006).
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    [CrossRef]
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    [CrossRef]
  21. S. Velghe, J. Primot, N. Guérineau, R. Haidar, S. Demoustier, M. Cohen and B. Wattellier, "Advanced wave-front sensing by quadri-wave lateral shearing interferometry," Proc. SPIE 6292, 62920E (2006).
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    [CrossRef] [PubMed]

2008 (2)

J. Zou, A. Sautivet, J. Fils, L. Martin, K. Abdeli, C. Sauteret and B. Wattellier, "Optimization of the dynamic wavefront control of a pulsed kilojoule/nanosecond-petawatt laser facility," Appl. Opt. 47(5), 704-710 (2008).
[CrossRef]

A. Sergé, N. Bertaux, H. Rigneault and D. Marguet, "Dynamic multiple-target tracing to probe spatiotemporal cartography of cell membranes," Nat. Methods 5, 687-695 (2008).
[CrossRef] [PubMed]

2007 (1)

W. Choi, C. Fang-Yen, K. Badizadegan, S. Oh, N. Lue, R. R. Dasari and M. S. Feld, "Tomographic phase microscopy," Nature Methods 4, 717-719 (2007).
[CrossRef] [PubMed]

2006 (4)

F. Charrière, A. Marian, F. Montfort, J. Kuehn, T. Colomb, E. Cuche, P. Marquet and C. Depeursinge, "Cell refractive index tomography by digital holographic microscopy," Opt. Lett. 31(2), 178-180 (2006).
[CrossRef]

C. Curl, C. J. Bellair, P. J. Harris, B. E. Allman, A. Roberts, K. A. Nugent and L. M. D. Delbridge, "Single cell volume measurement by Quantitative Phase Microscopy (QPM): A case study of Erythrocyte Morphology," Cellular Physiology Biochem. 17, 193-200 (2006).
[CrossRef] [PubMed]

S. Bernet, A. Jesacher, S. Fürhapter, C. Maurer and M. Ritsch-Marte, "Quantitative imaging of complex samples by spiral phase contrast microscopy," Opt. Express 14(9), 3792-3805 (2006).
[CrossRef]

S. Velghe, J. Primot, N. Guérineau, R. Haidar, S. Demoustier, M. Cohen and B. Wattellier, "Advanced wave-front sensing by quadri-wave lateral shearing interferometry," Proc. SPIE 6292, 62920E (2006).

2005 (1)

2004 (2)

M. R. Arnison, K. G. Larkin, C. J. R. Sheppard, N. I. Smith, and C. J. Cogswell, "Linear phase imaging using differential interference contrast microscopy," J. Microscopy 214(1), 7-12 (2004).
[CrossRef]

C. J. Bellair, C. L. Curl, B. E. Allman, P. J. Harris, A. Roberts and L. M. D. Delbridge, "Quantitative phase amplitude microscopy IV: imaging thick specimens," J. Microscopy 214(1), 62-68 (2004).
[CrossRef]

2003 (1)

J. Primot, "Theoretical description of Shack-Hartmann wave-front sensor," Opt. Commun. 222, 81-92 (2003).
[CrossRef]

2001 (1)

Y. Xu, Y. x. Xu, M. Hui and X. Cai, "Quantitative surface topography determination by differential interference contrast microscopy," Opt. Precision Eng. 9, 226-229 (2001).

2000 (2)

1999 (1)

1998 (1)

1996 (1)

1995 (1)

1976 (1)

C. S. Chong and C. Colbow, "Light scattering and turbidity measurements on lipid vesicles," Biochim. Biophys. Acta 436,260-282 (1976).
[CrossRef] [PubMed]

1971 (1)

R. V. Shack and B. C. Platt, "Production and use of a lenticular Hartmann screen," J. Opt. Soc. Am. 61, 656 (1971).

1964 (1)

F. C. Jensen, J. Anthony, R. V. G. Girardi and H. Koprowski (1964), "Infection of Human and Simian Tissue Cultures with Rous Sarcoma Virus," PNAS 52(1), 53-59 (1964).
[CrossRef]

1955 (1)

G. Nomarski, "Nouveau Dispositif Pour l’Observation En Contraste De Phase Differentiel," J. Phys. Radium 16, S88-S88 (1955).

1935 (1)

F. Zernike, "Das Phasenkontrastverfahren bei der mikroskopischen Beobachtung," Z. Tech. Phys. 16, 454-457 (1935).

Abdeli, K.

Allman, B. E.

C. Curl, C. J. Bellair, P. J. Harris, B. E. Allman, A. Roberts, K. A. Nugent and L. M. D. Delbridge, "Single cell volume measurement by Quantitative Phase Microscopy (QPM): A case study of Erythrocyte Morphology," Cellular Physiology Biochem. 17, 193-200 (2006).
[CrossRef] [PubMed]

C. J. Bellair, C. L. Curl, B. E. Allman, P. J. Harris, A. Roberts and L. M. D. Delbridge, "Quantitative phase amplitude microscopy IV: imaging thick specimens," J. Microscopy 214(1), 62-68 (2004).
[CrossRef]

Anthony, J.

F. C. Jensen, J. Anthony, R. V. G. Girardi and H. Koprowski (1964), "Infection of Human and Simian Tissue Cultures with Rous Sarcoma Virus," PNAS 52(1), 53-59 (1964).
[CrossRef]

Arnison, M. R.

M. R. Arnison, K. G. Larkin, C. J. R. Sheppard, N. I. Smith, and C. J. Cogswell, "Linear phase imaging using differential interference contrast microscopy," J. Microscopy 214(1), 7-12 (2004).
[CrossRef]

Badizadegan, K.

W. Choi, C. Fang-Yen, K. Badizadegan, S. Oh, N. Lue, R. R. Dasari and M. S. Feld, "Tomographic phase microscopy," Nature Methods 4, 717-719 (2007).
[CrossRef] [PubMed]

Barty, A.

Bellair, C. J.

C. Curl, C. J. Bellair, P. J. Harris, B. E. Allman, A. Roberts, K. A. Nugent and L. M. D. Delbridge, "Single cell volume measurement by Quantitative Phase Microscopy (QPM): A case study of Erythrocyte Morphology," Cellular Physiology Biochem. 17, 193-200 (2006).
[CrossRef] [PubMed]

C. J. Bellair, C. L. Curl, B. E. Allman, P. J. Harris, A. Roberts and L. M. D. Delbridge, "Quantitative phase amplitude microscopy IV: imaging thick specimens," J. Microscopy 214(1), 62-68 (2004).
[CrossRef]

Bernet, S.

Bertaux, N.

A. Sergé, N. Bertaux, H. Rigneault and D. Marguet, "Dynamic multiple-target tracing to probe spatiotemporal cartography of cell membranes," Nat. Methods 5, 687-695 (2008).
[CrossRef] [PubMed]

Bevilacqua, F.

Charrière, F.

Choi, W.

W. Choi, C. Fang-Yen, K. Badizadegan, S. Oh, N. Lue, R. R. Dasari and M. S. Feld, "Tomographic phase microscopy," Nature Methods 4, 717-719 (2007).
[CrossRef] [PubMed]

Chong, C. S.

C. S. Chong and C. Colbow, "Light scattering and turbidity measurements on lipid vesicles," Biochim. Biophys. Acta 436,260-282 (1976).
[CrossRef] [PubMed]

Cogswell, C. J.

M. R. Arnison, K. G. Larkin, C. J. R. Sheppard, N. I. Smith, and C. J. Cogswell, "Linear phase imaging using differential interference contrast microscopy," J. Microscopy 214(1), 7-12 (2004).
[CrossRef]

Cohen, M.

S. Velghe, J. Primot, N. Guérineau, R. Haidar, S. Demoustier, M. Cohen and B. Wattellier, "Advanced wave-front sensing by quadri-wave lateral shearing interferometry," Proc. SPIE 6292, 62920E (2006).

Colbow, C.

C. S. Chong and C. Colbow, "Light scattering and turbidity measurements on lipid vesicles," Biochim. Biophys. Acta 436,260-282 (1976).
[CrossRef] [PubMed]

Colomb, T.

Cuche, E.

Curl, C.

C. Curl, C. J. Bellair, P. J. Harris, B. E. Allman, A. Roberts, K. A. Nugent and L. M. D. Delbridge, "Single cell volume measurement by Quantitative Phase Microscopy (QPM): A case study of Erythrocyte Morphology," Cellular Physiology Biochem. 17, 193-200 (2006).
[CrossRef] [PubMed]

Curl, C. L.

C. J. Bellair, C. L. Curl, B. E. Allman, P. J. Harris, A. Roberts and L. M. D. Delbridge, "Quantitative phase amplitude microscopy IV: imaging thick specimens," J. Microscopy 214(1), 62-68 (2004).
[CrossRef]

Dasari, R. R.

W. Choi, C. Fang-Yen, K. Badizadegan, S. Oh, N. Lue, R. R. Dasari and M. S. Feld, "Tomographic phase microscopy," Nature Methods 4, 717-719 (2007).
[CrossRef] [PubMed]

Delbridge, L. M. D.

C. Curl, C. J. Bellair, P. J. Harris, B. E. Allman, A. Roberts, K. A. Nugent and L. M. D. Delbridge, "Single cell volume measurement by Quantitative Phase Microscopy (QPM): A case study of Erythrocyte Morphology," Cellular Physiology Biochem. 17, 193-200 (2006).
[CrossRef] [PubMed]

C. J. Bellair, C. L. Curl, B. E. Allman, P. J. Harris, A. Roberts and L. M. D. Delbridge, "Quantitative phase amplitude microscopy IV: imaging thick specimens," J. Microscopy 214(1), 62-68 (2004).
[CrossRef]

Demoustier, S.

S. Velghe, J. Primot, N. Guérineau, R. Haidar, S. Demoustier, M. Cohen and B. Wattellier, "Advanced wave-front sensing by quadri-wave lateral shearing interferometry," Proc. SPIE 6292, 62920E (2006).

Depeursinge, C.

Emery, Y.

Fang-Yen, C.

W. Choi, C. Fang-Yen, K. Badizadegan, S. Oh, N. Lue, R. R. Dasari and M. S. Feld, "Tomographic phase microscopy," Nature Methods 4, 717-719 (2007).
[CrossRef] [PubMed]

Feld, M. S.

W. Choi, C. Fang-Yen, K. Badizadegan, S. Oh, N. Lue, R. R. Dasari and M. S. Feld, "Tomographic phase microscopy," Nature Methods 4, 717-719 (2007).
[CrossRef] [PubMed]

Fils, J.

Fürhapter, S.

Girardi, R. V. G.

F. C. Jensen, J. Anthony, R. V. G. Girardi and H. Koprowski (1964), "Infection of Human and Simian Tissue Cultures with Rous Sarcoma Virus," PNAS 52(1), 53-59 (1964).
[CrossRef]

Guérineau, N.

S. Velghe, J. Primot, N. Guérineau, R. Haidar, S. Demoustier, M. Cohen and B. Wattellier, "Advanced wave-front sensing by quadri-wave lateral shearing interferometry," Proc. SPIE 6292, 62920E (2006).

J. Primot and N. Guérineau, "Extended Hartmann Test Based on the Pseudoguiding Property of a Hartmann Mask Completed by a Phase Chessboard," Appl. Opt. 39(31), 5715-5720 (2000).
[CrossRef]

Haidar, R.

S. Velghe, J. Primot, N. Guérineau, R. Haidar, S. Demoustier, M. Cohen and B. Wattellier, "Advanced wave-front sensing by quadri-wave lateral shearing interferometry," Proc. SPIE 6292, 62920E (2006).

Hariharan, P.

Harris, P. J.

C. Curl, C. J. Bellair, P. J. Harris, B. E. Allman, A. Roberts, K. A. Nugent and L. M. D. Delbridge, "Single cell volume measurement by Quantitative Phase Microscopy (QPM): A case study of Erythrocyte Morphology," Cellular Physiology Biochem. 17, 193-200 (2006).
[CrossRef] [PubMed]

C. J. Bellair, C. L. Curl, B. E. Allman, P. J. Harris, A. Roberts and L. M. D. Delbridge, "Quantitative phase amplitude microscopy IV: imaging thick specimens," J. Microscopy 214(1), 62-68 (2004).
[CrossRef]

Jensen, F. C.

F. C. Jensen, J. Anthony, R. V. G. Girardi and H. Koprowski (1964), "Infection of Human and Simian Tissue Cultures with Rous Sarcoma Virus," PNAS 52(1), 53-59 (1964).
[CrossRef]

Jesacher, A.

Koprowski, H.

F. C. Jensen, J. Anthony, R. V. G. Girardi and H. Koprowski (1964), "Infection of Human and Simian Tissue Cultures with Rous Sarcoma Virus," PNAS 52(1), 53-59 (1964).
[CrossRef]

Kuehn, J.

Larkin, K. G.

M. R. Arnison, K. G. Larkin, C. J. R. Sheppard, N. I. Smith, and C. J. Cogswell, "Linear phase imaging using differential interference contrast microscopy," J. Microscopy 214(1), 7-12 (2004).
[CrossRef]

Lue, N.

W. Choi, C. Fang-Yen, K. Badizadegan, S. Oh, N. Lue, R. R. Dasari and M. S. Feld, "Tomographic phase microscopy," Nature Methods 4, 717-719 (2007).
[CrossRef] [PubMed]

Magistretti, P. J.

Marguet, D.

A. Sergé, N. Bertaux, H. Rigneault and D. Marguet, "Dynamic multiple-target tracing to probe spatiotemporal cartography of cell membranes," Nat. Methods 5, 687-695 (2008).
[CrossRef] [PubMed]

Marian, A.

Marquet, P.

Martin, L.

Maurer, C.

Montfort, F.

Nagle, J. F.

J. F. Nagle and S. Tristram-Nagle, "Structure of lipid bilayers," Biochim. Biophys. Acta 1469(3), 159-195 (2000).

Nomarski, G.

G. Nomarski, "Nouveau Dispositif Pour l’Observation En Contraste De Phase Differentiel," J. Phys. Radium 16, S88-S88 (1955).

Nugent, K. A.

C. Curl, C. J. Bellair, P. J. Harris, B. E. Allman, A. Roberts, K. A. Nugent and L. M. D. Delbridge, "Single cell volume measurement by Quantitative Phase Microscopy (QPM): A case study of Erythrocyte Morphology," Cellular Physiology Biochem. 17, 193-200 (2006).
[CrossRef] [PubMed]

A. Barty, K. A. Nugent, D. Paganin and A. Roberts, "Quantitative optical phase microscopy," Opt. Lett. 23(11), 817-819 (1998).
[CrossRef]

Oh, S.

W. Choi, C. Fang-Yen, K. Badizadegan, S. Oh, N. Lue, R. R. Dasari and M. S. Feld, "Tomographic phase microscopy," Nature Methods 4, 717-719 (2007).
[CrossRef] [PubMed]

Paganin, D.

Platt, B. C.

R. V. Shack and B. C. Platt, "Production and use of a lenticular Hartmann screen," J. Opt. Soc. Am. 61, 656 (1971).

Primot, J.

S. Velghe, J. Primot, N. Guérineau, R. Haidar, S. Demoustier, M. Cohen and B. Wattellier, "Advanced wave-front sensing by quadri-wave lateral shearing interferometry," Proc. SPIE 6292, 62920E (2006).

J. Primot, "Theoretical description of Shack-Hartmann wave-front sensor," Opt. Commun. 222, 81-92 (2003).
[CrossRef]

J. Primot and N. Guérineau, "Extended Hartmann Test Based on the Pseudoguiding Property of a Hartmann Mask Completed by a Phase Chessboard," Appl. Opt. 39(31), 5715-5720 (2000).
[CrossRef]

J. Primot and L. Sogno, "Achromatic three-wave (or more) lateral shearing interferometer," J. Opt. Soc. Am. A 12(12), 2679 (1995).
[CrossRef]

Rappaz, B.

Rigneault, H.

A. Sergé, N. Bertaux, H. Rigneault and D. Marguet, "Dynamic multiple-target tracing to probe spatiotemporal cartography of cell membranes," Nat. Methods 5, 687-695 (2008).
[CrossRef] [PubMed]

Ritsch-Marte, M.

Roberts, A.

C. Curl, C. J. Bellair, P. J. Harris, B. E. Allman, A. Roberts, K. A. Nugent and L. M. D. Delbridge, "Single cell volume measurement by Quantitative Phase Microscopy (QPM): A case study of Erythrocyte Morphology," Cellular Physiology Biochem. 17, 193-200 (2006).
[CrossRef] [PubMed]

C. J. Bellair, C. L. Curl, B. E. Allman, P. J. Harris, A. Roberts and L. M. D. Delbridge, "Quantitative phase amplitude microscopy IV: imaging thick specimens," J. Microscopy 214(1), 62-68 (2004).
[CrossRef]

A. Barty, K. A. Nugent, D. Paganin and A. Roberts, "Quantitative optical phase microscopy," Opt. Lett. 23(11), 817-819 (1998).
[CrossRef]

Sauteret, C.

Sautivet, A.

Sergé, A.

A. Sergé, N. Bertaux, H. Rigneault and D. Marguet, "Dynamic multiple-target tracing to probe spatiotemporal cartography of cell membranes," Nat. Methods 5, 687-695 (2008).
[CrossRef] [PubMed]

Shack, R. V.

R. V. Shack and B. C. Platt, "Production and use of a lenticular Hartmann screen," J. Opt. Soc. Am. 61, 656 (1971).

Sheppard, C. J. R.

M. R. Arnison, K. G. Larkin, C. J. R. Sheppard, N. I. Smith, and C. J. Cogswell, "Linear phase imaging using differential interference contrast microscopy," J. Microscopy 214(1), 7-12 (2004).
[CrossRef]

Smith, N. I.

M. R. Arnison, K. G. Larkin, C. J. R. Sheppard, N. I. Smith, and C. J. Cogswell, "Linear phase imaging using differential interference contrast microscopy," J. Microscopy 214(1), 7-12 (2004).
[CrossRef]

Sogno, L.

Tristram-Nagle, S.

J. F. Nagle and S. Tristram-Nagle, "Structure of lipid bilayers," Biochim. Biophys. Acta 1469(3), 159-195 (2000).

Velghe, S.

S. Velghe, J. Primot, N. Guérineau, R. Haidar, S. Demoustier, M. Cohen and B. Wattellier, "Advanced wave-front sensing by quadri-wave lateral shearing interferometry," Proc. SPIE 6292, 62920E (2006).

Wattellier, B.

J. Zou, A. Sautivet, J. Fils, L. Martin, K. Abdeli, C. Sauteret and B. Wattellier, "Optimization of the dynamic wavefront control of a pulsed kilojoule/nanosecond-petawatt laser facility," Appl. Opt. 47(5), 704-710 (2008).
[CrossRef]

S. Velghe, J. Primot, N. Guérineau, R. Haidar, S. Demoustier, M. Cohen and B. Wattellier, "Advanced wave-front sensing by quadri-wave lateral shearing interferometry," Proc. SPIE 6292, 62920E (2006).

Xu, Y.

Y. Xu, Y. x. Xu, M. Hui and X. Cai, "Quantitative surface topography determination by differential interference contrast microscopy," Opt. Precision Eng. 9, 226-229 (2001).

Zernike, F.

F. Zernike, "Das Phasenkontrastverfahren bei der mikroskopischen Beobachtung," Z. Tech. Phys. 16, 454-457 (1935).

Zou, J.

Appl. Opt. (3)

Biochim. Biophys. Acta (2)

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Supplementary Material (2)

» Media 1: AVI (14264 KB)     
» Media 2: AVI (3890 KB)     

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Figures (9)

Fig. 1.
Fig. 1.

Schematic layout of the setup.

Fig. 2.
Fig. 2.

(a) Ideal transparency for a 4-wave-only 2D diffraction grating. (b) MHM transparency, approximation of (a). (c) Intensity transmission of (a). (d) 1D interferogram formation by a MHM in the case of a disturbed input wavefront; geometrical approach. (e) 2D interferogram formation by a MHM in the case of a plane input wavefront; visualization of the 4 diffracted waves (arrows).

Fig. 3.
Fig. 3.

(a) Interferogram obtained with an aberrant wavefront. (b) Zoom on a part of (a), visualization of the local frequency modulations. (c) Fourier Transform of (a). (d) Obtaining of 2 OPD orthogonal gradients by Inverse Fourier Transform of 2 sub-images of (b). (e) 2D Integration of the gradients to reconstruct the OPD.

Fig. 4.
Fig. 4.

Principle of the interferogram formation in case of an incoherent illumination.

Fig. 5.
Fig. 5.

Diminution of the sensitivity according to the spatial coherence of the source.

Fig. 6.
Fig. 6.

Contrast comparison of simulated interferograms and acquired interferograms, for different input illumination numerical apertures. Obtained with z=1420mm, p=29.6µm and gy =20.

Fig. 7.
Fig. 7.

(a) Intensity image of a 5µm polystyrene bead, in imaging conditions with SID4-HR, into a n=1.552 transparent liquid hydrocarbon ; (b) Phase measurement (false colors) ; (c) 3D phase visualization (false colors); (d) Phase shift profile along the center of the bead, comparison between measurement and model.

Fig. 8.
Fig. 8.

(a) Intensity image, in imaging conditions, of a DPPC GUV without optical index difference between inside and outside the GUV ; (b) Phase-shift image ; (c) Phase-shift value compared to the distance from the GUV center. Red squares : Azimuthal average of the phase-shift map; the symmetry before and after x=0 is explained by the complete 2π averaging. Vertical red lines : error bars, Azimuthal standard deviation. Solid black line : Theoretical phase-shift profile obtained for D=16.65µm, e=3.85nm and δn=nDPPC -nmedium =0.128.

Fig. 9.
Fig. 9.

Single frame excerpts from (Media 1), (Media 2). (a) Intensity image, in imaging conditions, of COS-7 cells ; (b) Phase image.

Equations (20)

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A (r)=I(r)exp(i[k·rφ(r)])
φ(r)=2πλOPD+φ(r0)
t (x,y)=cos(πxp) · c o s (πyp)
T(x,y)=t(x,y)2=14{1+[cos(2πpx)+cos(2πpy)]+12[cos(2πp(x+y))+cos(2πp(xy))]}
I(r,z)=I0{1+[cos(2πpx+2πpzOPDx)+cos(2πpy+2πpzOPDy)]+12[cos(2πp(x+y)+2πpzOPD(x+y))+cos(2πp(xy)+2πpzOPD(xy))]}
δ=z·OPD
HxHy=2πpzOPDxOPDy
I(x,y)= Sd2I(x,y,xs,ys)
d2 I(x,y,xs,ys)=Iponc(x+xs,y+ys)S·dxsdys
Iponct(x,y)=I0· [1+cos(Kx)+cos(Ky)+12cos(2Kx+y2)+12cos(2Kxy2)]
I(x,y)=ϕs=02πrs=0RIponct(x+rs·cos(ϕs),y+rs·sin(ϕs))πR2·rsdrsdϕs
I(x,y)=I0[1+M(KR)[cos(Kx)+cos(Ky)]+12M(2KR)[cos(K[x+y])+cos(K[xy])]]
M(X)=2J1(X)X
R(D)=z·tan(θo)gy×11z·DfTL2=Rafocal×11z·DfTL2
SNRrelative(KR)=M(KR)ImaxI0
Imax,minI0 {1+2·M(KR)+M(2KR)1M(2KR)12·M(KR)+M(2KR)
r=0.61×λNAobj
4·a=pr·gy
C=max(I)min(I)max(I)+min(I)=ImaxIminImax+Imin
δ(x)={φmaxcos[arcsin(2xD)]ifx<D20else

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