Masters Thesis Defense - Development of Automated Brightfield Imaging for Cryoplane Microscopy
Date: December 22, 2005
Time: 11:00 AM
Location: Queen Lane Campus, Room: 261
Advisor: Jonathan Nissanov, Ph.D.
Cryoplane Microscope (CM) is a device used to examine biological material at a macro level of magnification using brightfield imaging and at a macro/micro level of magnification using epifluorescence microscopy. On this system, a frozen tissue is cut using a cryostat and the exposed blockface surface is imaged macroscopically using a line scan camera triggered by a linear encoder and then by an area scan camera attached to a microscope for micro level resolution. The tissue moves horizontally in our CM while the knife descends vertically. All imaging devices are mounted on the knife housing and descended along with the knife and thus focus is maintained on the blockface surface. By cutting the tissue block, deeper tissue levels are revealed. Successive planes are obtained as a series to yield an aligned dataset.
A Virtual Instrument (VI) using LabVIEW was developed as part of this thesis. It includes subVIs for monitoring tissue position, controlling intensity of brightfield illumination and image acquisition. Each line of the line scanner was triggered by the VI using data from a linear encoder monitoring tissue position. The pitch of the acquired line scan images is 14 Ám and those provide anatomical context for the higher resolution, up to 0.7 Ám pitch, obtained with the epifluorescence microscopy.
CM is being used in a variety of applications. Two notable ones are fluorescent imaging of gene expression using GFP transgenics and mapping of neural connectivity using fluorescence tract tracers. In both settings, the developed brightfield imaging system in conjunction with developed systemic stains will provide the anatomical framework to aid in understanding of the fluorescent signal obtained with the epifluorescent microscope.
The Queen Lane Campus is located at 2900 Queen Lane, Philadelphia, PA. A shuttle is available from 33rd and Market Streets