digital holography microscopy
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DIGITAL HOLOGRAPHICMICROSCOPY
PRESENTED BY : UNDER THE
RIMPLE POONIA GUIDANCE OF:
2010PHA2765 PROF. JOBY JOSEPH
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Reconstruction methods:
Fresnel approach: diffraction at anyaperture given by Fresnel Kirchoff integral
,
, (,)
( )( )Using approximation : and values, and values small compared to d we have for real
image
2
2
18
) 3 . .
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For Fourth term
)+
2
2
Using in above equation
, exp i2
exp
x,y h x,y exp i exp 2
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One fourier transform involved
Minimum distance requirement Pixel distance in reconstructed images varies with the
reconstruction distance as
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Convolution approach:
Numerical processing of Fresnel Kirchhoff integral is timeconsuming
Reconstruction formula with superposition integral
, , , (, , , ) With impulse response
, , , + + + +
System characterised by , , , , Convolution theorem is applied
Three Fourier transforms involved:
, . .
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Digital holographic microscopy
Lateral resolution of holographic reconstruction is given
by
[1
]
where =distance between source point and hologramplane in recording process
=distance between source point and hologram plane inreconstruction process
and wavelenghts during recording and reconstructionFor desired magnification reconstruction distance
. and
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Source point needs to be
placed at
[ 1 1
1]
Reference wave needs to be
placed at
, exp( 2
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Fresnel
approach
Convolution
approach
d=37cm
Reconstructions
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Fresnel
approach
Convolution
approach
d=37cm
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Interesting Property
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Intensity at the hologram plane
Reconstructed wavefront at hologram plane
Zero order removal
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Average intensity subtraction
Average intensity of all pixels in hologram
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High pass filtering-steps involved
Read the hologram
Convert from spatial domain to frequency domain with FFT2 and
shift the zero components with FFTSHIFT
Define a circular mask and position it at the center of the Fourier
transformed image
Return to spatial domain with IFFT2 and IFFTSHIFT2
Save as tiff image
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High pass filtering
Average subtraction
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Spatial filtering-steps involved
Read the hologram
Convert from spatial domain to frequency domain with FFT2 and shift
the zero components with FFTSHIFT
Define masks numerically according to the spectrum of the hologram
and multiply with the Fourier transform obtained above
Return to spatial domain with IFFT2 and IFFTSHIFT2
Filtered hologram is obtained which is numerically reconstructed
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Spatial filtering
Spectrum of the hologram
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Twin image elimination
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FIR METHOD
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Autofocusing evaluation functions
In autofocusing determination and maximization of image
contrast needed
For amplitude objects-focused image at maximal focusvalue
For phase objects-minimum absorption-focused image at
minimal focus value Evaluation functions:
Variance of the grey function
based on the statistical analysis of the gray value
distribution Sharp structures in a focused image result in a higher
contrast than in a smooth defocused image
1 ,
=
=
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Weighted Fourier spectral analysis
quantifies the sharpness of the edges as a focused image
contains more fine details than defocused image
log[1(,)]== Standard deviation correlation functionThereconstructed image field is given by
, , (,,0)
And intensity
I , , , z ( , , , z)
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laser
Pin
hole
lens
Beam splitter1NF
mirror
mirror
object
MO
Beam
splitter2
CCD
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Result
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At d=30 cm
At d=45 cm At d=60cm
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REFERENCES B. Kemper, and G. Bally, "Digital holographic microscopy for live cell applications and
technical inspection," Appl.Opt. 47(2), A52-A61 (2008).
Collier R. J., Burckhardt C. B., and Lin L.H., Optical Holography, (Academic Press,
New York, 1971).
Christopher J Mann, L Yu and K Kim* Movies of cellular and sub-cellular motion by
digital holographic microscopy, (BioMedical Engineering OnLine2006, 5:21)
Cuche E., Marquet P., and Depeursinge C., Spatial filtering for zero-order and twin
image elimination in digital off axis holography, Appl.Opt.39 (2000) 4070-4075.
Cuche E, Marquet P, and Depeursinge C,Simultaneous amplitude-contrast and
quantitative phase-contrast microscopy by numerical reconstruction of Fresnel off-axis
holograms,( Appl. Opt , Vol. 38, 1999)
Dubois F, Schockaert C,Callens N and Yourassowsky C,Focus plane detection
criteria in digital holography microscopy by amplitude analysis (Optical Society of
America, 2006) Gabor D, A new microscope principle, Nature, 161 (1948) 777-778.
Goodman J. W, Introduction to Fourier Optics, (McGraw-Hill, New York, 1996).
Hariharan P., Optical Holography,:Principle, techniques, and applications, (Cambridge
University Press, Cambridge ,1983).
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Sucerquia J, Xu W, Stephan K. Jericho,Klages P,Manfred H. Jericho, and H. Jrgen Kreuzer, Digital in-line
holographic microscopy (Appl. Opt Vol. 45, 2006)
Kreis Th, Adams M., and JptnerW., Methods of digital holography: a comparison, Proc. SPIE 3098 (1997) 224
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Kreis T.M., and JuptnerW., Suppression of dc term in digital holography, Opt.Engn. 36 (1997) 2367-2370.
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Liu W,Kang X , Xiaoyuan He .Filter method used to eliminate the zero-order image,( SPIE Vol. 7375, 2009) Lohmann A. W., Mendlovic David, and Shabtay Gal, Significance of phase and amplitude in the Fourier domain, J.
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M. Liebling, M. Unser, Autofocus for digital Fresnel holograms by use of a Fresnelet-sparsity criterion,( J. Opt.Soc.
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M. K. Kim, Principles and techniques of digital holographic microscopy(SPIE Rev. 1 2010.
P. Marquet, B. Rappaz, P. J. Magistretti, E. Cuche, Y. Emery, T. Colomb, C. Depeursinge,Digital holographic
microscopy: a non invasive contrast imaging technique allowing quantitative visualization of living cells with sub
wavelength accuracy, (Opt. Lett. 30, 468 -470,2005)
Schnars U. and JuptnerW. Direct recording of holograms by a CCD target and numerical reconstruction, Appl. Opt.33 (1994) 79181.
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Wang H, Aili Qin, Min Huang,Autofocus method for digital holographic reconstruction of microscopic object,.(
IEEE,2009)
Wang H, Aili Qin, Min et al,Autofocus method for amplitude and pure phase objects reconstruction in digital
holography.( Proc. of SPIE Vol. 7513,2009)
Wang H , Lihong M Yong Li,Haijun Zhang, Elimination of zero-order diffraction and conjugate image in off-axis digital
holography (Journal of Modern Optics, 56:21, 2377-2383, 2009)
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