Life Sciences Commons^™

Light Microscopy Reflection Focusing Cells And Coverglass, George Mcnamara

George McNamara

Light Microscopy Reflection Focusing Cells and Coverglass 20150520Wed

Many light microscopes have the capability of using camera based brightfield, phase contrast, or fluorescence to find the most contrasty specimen focus. When I worked for UIC (1992-1997) we offered this with a combination of Ludl MAC2000 controller, Z-drive, video card thingy, and video camera (most customers at the time used analog video cameras).

Many automated light microscopes now have clever -- and pricey -- autofocus systems based on an NIR LED structured illumination reflecting light off the bottom of the coverglass.

I encourage a simpler approach - my hardware details are …

Image Z-Series Wrt Nvidia Titan 6 Gb Gpu Cuda Deconvolution, George Mcnamara

George McNamara

Image Z-series wrt NVidia Titan 6 Gb GPU CUDA Deconvolution

This ZIP file contains raw and deconvolved data series from George McNamara, Laurence J.N. Cooper lab, M.D. Anderson Cancer Center. I have also included our lab instructions, and a screenshot of the current CUDA deconvolution settings (for dry objective lenses, Manish Butte wrote me to use the immersion medium refractive index, air = 1.0).

Instrument:

Leica DMI6000 inverted fluorescence microscope, Lumencor SOLA LED light source, Leica "L5" GFP filter cube (480/40ex, 505dm, 527/30em).

Leica 20x/0.75 NA objective lens, 1.6x optovar, for 325 nm XY piel size. For this dataset, I …

3b Project Data - Raw Tiff Series And Cropped Series, George Mcnamara

George McNamara

Fluorescence microscopy data acquired for 3B Bayesian localization microscopy.

Settings: 100 nm XY pixel size. 50 millisecond exposures, MetaMorph stream acquisition mode (as fast as possible). Stellaris FISH mammalian cells labeled with Quasar 670 fluorophores, one molecule per 20mer FISH probe, ~48 probes to TOP1 mRNA (Topoisomerase 1).

More on the 3B project and Stellaris FISH data at http://works.bepress.com/gmcnamara/38/

and at Susan Cox's web sites http://www.coxphysics.com/3b/ http://superresolved.com/

Leica Microscope Gpu Deconvolution Stellaris Fish Dataset #1, George Mcnamara

George McNamara

McNamara 20131101Fri Leica widefield microscope CUDA Deconvolution Stellaris FISH probe cultured cells dataset #1.zip

A text file in the zip archive has experiment details. I am posting this online so that researchers - whether academic or commercial - can evaluate the acquired data, the Bruce and Butte 2013 Optics Express ( http://www.opticsinfobase.org/oe/fulltext.cfm?uri=oe-21-4-4766 ) deconvolution result (note: I may not have used optimal settings), and to compare these deconvolution results to other methods. If anyone generates alternative spatial deconvolution output, such as from: * SVI Huygens * Media Cy AutoQuant * Agard's ER-Decon (Arigovindan 2013 PNAS) * Vicidomini SGP GPU Deconv …

Flash4 Dark Reference Images, George Mcnamara

George McNamara

Hamamatsu FLASH4.0 dark reference images, acquired with 10 second exposure times, no light to camera. Camera offset (set by Hamamatsu( is ~100 (the average intensity of the first image is always ~1 intensity level higher - an odd feature, but trivial in practice for a 16-bit camera).

George McNamara, Ph.D.

Single Cells Analyst at L.J.N. Cooper Lab

University of Texas M.D. Anderson Cancer Center

Mcnamara 2011 Mpmicro - Multi-Probe Microscopy (10/31/2011), George Mcnamara

George McNamara

Multi-Probe Microscopy is an ~1500 page Word document summarizing what I know and/or found interesting in light microscopy, fluorescence microscopy and digital image analysis, from 1995-2005. Very little has been updated since 2005.