Holographic Video Microscopy
 
 
Thanks for your interest in our group's holographic video microscopy techniques.  Please visit our tutorial page for using our holographic microscopy software.  
 
 
In the meanwhile, here are some general pointers to frequently asked queries regarding this technique.
 
Q1: I am working on 3D tracking of particles. I get  hologram but facing some difficulty in computation. I want to do the following things.
 
1- reconstruct the normalize hologram and numerical fit of B(p).
 
2- then trace the particle in 3D.
 
 
General pointers:
1. The idea of normalization is to eliminate the effect of nonuniform illumination
by dividing the hologram of your sample by a "background" hologram.  Typically,
we obtain background holograms in two ways: (a) taking a holographic snapshot
with none of the sample particles in the field of view or (b) taking lots of images
of the field of view with particles moving about vigorously, and computing the
median image of all of those images.
 
Once you have the background hologram, you normalize by dividing each holographic
snapshot by the background.  You have to take care that the background has no
zero-value pixels, but that's about it.
 
After normalization, the mean value of the image should be around 1.
 
Please refer to the tutorial for more help on using spheretool.pro
 
2. The IDL routine called SPHERETOOL is supposed to provide a gentle
introduction to the whole fitting process.  It calls all of the holographic microscopy
software and some of our utility routines.  It also relies on the MPFIT software
distributed by Craig Markwardt and (optionally) the GPUlib routines for
hardware-accelerated computation.
 
Once you have everything installed, you use it as
IDL> spheretool, float(a) / (background > 1)
 
The float() call avoids rounding the pixel values to integers.  The ">" ensures that
zero-valued pixels in the background won't cause problems.
 
The code also has comments explaining what everything is and what it does.
 
 
Specific pointers:
 
This image looks like the illuminating beam was not collimated.  Did you illuminate
through the microscope's condenser lens?  If so, don't.  Move the condenser out of way,
and fire a collimated laser beam straight down the optical axis through the sample and
into the objective lens.  The problem is that the spheres' images are distorted into
ellipses by an amount that depends on their distance from the center of the field
of view.  Accounting for this in software will be __VERY__ difficult.  Fixing it
with proper optical alignment is much much easier.
 
 
Please feel free to write to us if you have more queries.
 
Contact us
  1.     David G. Grier: david.grier@nyu.edu
  2.     Fook Chiong Cheong: phycfc@gmail.com