This dataset was acquired to test the accuracy of image stitching
tools. It is acquired by imaging stem cell colonies seeded in one
well of a 6-well plate using in phase contrast and fluorescent
modalities. All plates are seeded at low density to allow for
colony growth. They are then fixed at days 2, 3, and 4 respectively.
Reference imaging measurements
The exposure time of the Cy5 fluorescent imaging modality was
selected to produce high contrast images where a manual
segmentation threshold of 500 intensity unit (pixels of intensity
greater than 500 are foreground) correctly segments an image.
Each of the three reference datasets consist of a pool of centered
reference colony images and their associated stage locations. The
area and centroid locations are acquired after segmenting raw
images with a global manually selected threshold and filtering
small objects by imposing a minimum colony size of 3 stem cells
(area > 2000 pixels). Each colony is centered inside the FOV by
an automated feedback loop providing translations to geometric
centroid of a thresholded segmented mask of that colony. The loop
minimizes the difference between the colony centroid and the middle
of the FOV and a convergence is reached when that difference is
less than 1 μm. The final centroid location for a colony is saved
from the microscope stage controller. The three reference datasets
(Day2, Day3, and Day4) were acquired using an Olympus microscope
controlled by MicroManager.
Dataset for testing stitching accuracy
For each well of the 3 plates, five sets of image tiles were
acquired. Table 1 summarizes these datasets. Each set of image
tiles covers the same inscribed square region on the plate as
the reference measurements. This test dataset is imaged with
overlaps ranging from 10 % to 50 %. The variation in the overlap
between image tiles enables the evaluation of stitching algorithms
with respect to the overlap.