Biomedical Engineering Reference
In-Depth Information
to OPD
m
and not for direct imaging. As long as the amplified noise does not exceed
λ
m
/2,
this process remains valid and the single-wavelength noise level
ε
m
can be maintained
while the unambiguous measurement range can be increased to
12
. Since two-wavelength
unwrapping relies on a mathematical analytical solution rather than the gradient
minimization strategies, this unwrapping method is particularly useful for structures with
sharp phase discontinuities.
Λ
There is an inherent trade-off between measurement range and noise amplification in two-
wavelength phase unwrapping. As the separation between
λ
1
and
λ
2
is decreased, both the
synthetic wavelength,
12
, and the total measurement range increase. With unlimited
signal-to-noise ratio and perfect system stability, the measurement range could be extended
arbitrarily using two laser sources with an infinitesimally small wavelength spacing;
however, practical noise limits require careful instrument design and illumination selection
in order to extend the measurement range while avoiding overamplification of the phase
noise. To address this limitation of noise amplification, Mann et al.
[37]
proposed a
hierarchical method of phase unwrapping that uses three wavelengths and intermediate
synthetic wavelength profiles.
Λ
In order to use multiple-wavelength phase unwrapping with dynamic samples, detection and
illumination schemes that allow the acquisition of multiple measurements in a single
snapshot are required. Previously, reflection-geometry holographic microscopy systems
have used spatial frequency multiplexing
[37,38]
or camera color channels with matched
illumination wavelengths
[39,40]
to simultaneously acquire separate complex wavefront
information. However, transmission-geometry phase measurements for semitransparent
samples had only been demonstrated with sequential illumination schemes that are intended
for measuring static samples
[41,42]
.
In order to adapt multiple-wavelength unwrapping to imaging of dynamic semitransparent
samples, such as cell cultures and other biological phenomena in microfluidic devices, we
proposed the use of an RGB color camera with transmission-geometry QPM
[43]
. The
illumination of the optical system shown in
Figure 14.6A
consisted of two lasers (
532
and 633 nm) that were chosen to match the peak spectral responses of the
red
and
green
color channels of the Bayer pattern color camera (Roper Scientific, CoolSNAP
cf
) as seen
in
Figure 14.6B
. Light from the two interferometer arms was collected by matched MO and
imaged onto the camera by common tube lens L
1
. A slight lateral shift in the reference arm
MO created a linear off-axis interference fringe for each illumination wavelength.
λ5
Off-axis interferograms were obtained simultaneously for each wavelength. Because the
color channels have low-intensity crosstalk, 4.3% and 5.4% for the green and red channels,
respectively, the interferograms were easily isolated. After separating the color channels,
the complex information in the Fourier domain was spatially filtered and recentered
according to the methods of Cuche et al.
[44]
. This operation removed the zero-order
Search WWH ::
Custom Search