3-D Microscopy (Deconvolution and Confocal)

Location: Thomas Building, DE-512
Contact phone: (206) 667-4205
Contact e-mail: imaging@fredhutch.org

Deconvolution and confocal microscopy are optical sectioning techniques for capturing high resolution 2-D or 3-D images of biological specimens. Wide field deconvolution uses conventional widefield microscopy and deconvolution software to remove out of focus blur.


Fluorescently labeled cells imaged with conventional widefield fluorescence microscopy without (left) and with deconvolution (right), showing the effect of deconvolution in removing out of focus haze and improving resolution and contrast.


Effect of Deconvolution
Deconvolution (right) removes out of focus haze and improves resolution and contrast.

This method provides superb results with thin specimens such as bacteria, yeasts, cell monolayers, thin sections, and other samples no more than 10-15 microns thick. Confocal microscopy uses optical techniques to remove out of focus haze and capture high contrast images of in-focus planes. This method is recommended for thicker specimens such as tissue sections, fly embryos, embryonic and adult worms, isolated tissues, cell spheroids, and other specimens. Several confocal technologies are available, including laser scanning confocal microscopy (i.e. conventional confocal), spinning disk, and swept field confocal microscopy.

Equipment:

(for technical details, follow the links to individual microscopes)

GE/Applied Precision Deltavison Elite image restoration (deconvolution) microscope. Specially suited for high resolution 3-D imaging of thin samples (bacteria, yeasts, cell monolayers, some thin sections).

Zeiss LSM 780 NLO spectral laser scanning confocal and multi photon microscope. Ideal for the imaging of thick specimens (worm embryos, adult worms, fly embryos, tissue sections, biopsies, cell spheroids, and more).

Perkin Elmer Ultraview spinning disk confocal system. Specially recommended for time lapse experiments with live cells or small model systems (such as worm and fly embryos).

Nikon Ti Live swept field confocal. Primarily used for widefield fluorescence imaging, but can be used in confocal mode. Suitable for imaging CFP/YFP samples.

Scheduling time for instruments, training, and support:

To schedule time on any of our microscopes, or to schedule staff assistance or training, please use iLab. The resource is open to all from 9:00 AM - 5:00 PM, Monday through Friday. After hours access can be granted to experienced users and requires training by the resource's staff. Please contact Scientific Imaging for further details.

Please notify the resource if you need to cancel or change a reservation, so that we can make instruments available to others if needed. Users who are more than one hour late without notice may lose their reservation.

How to access your data:

Data acquired on Scientific Imaging instruments is transferred to the si folder in the user's fred account. If you do not have a fred account, please submit the computing account application form. Once you have obtained a fred account, please contact Scientific Imaging so that we can create an si link for you. External users without access to fred can obtain their data via ftp, or can download to their own data storage device from one of our dedicated computers. Please note that no portable drives are allowed on instrument computers.