Nikon PCM2000 Confocal Microscope
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The Departmental Confocal System is based on a Nikon E800 microscope with epifluorescence and DIC capabilities. The objectives are 4x, 10x, 20x, 40x, 60x and 100x with the last three being oil immersion. |
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The confocal portion is a computer-controlled scan head using light from up to three lasers. The three detectors allow three-color imaging or two colors and DIC. |
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argon laser emits light in the blue spectrum at 457 or 488 nanometers
(nm) and is suitable for fluorescent dyes such as CFP, FITC, GFP
and Alexa 488. The Green HeNe laser emits at 543 nm and is suitable
for use with red dyes such as TRITC, Cy3 and Texas Red. The Red
HeNe emits at 633nm and should be used for far red dyes such as
Cy5 and Alexa 633.
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The lasers are different not only in emission spectra but in power. This can result in large differences in the apparent brightness of different fluorescent dyes. To optimize multichannel (multicolor) imaging, the output of individual dyes must be modified so that the output from all dyes is equalized. This is accomplished through judicious use of the neutral density filters in front of the argon and red HeNe lasers.
For help in recognizing which dyes are appropriate for this system, go to the spectra page and explore the realtionship between the spectral qualities of fluorescent dyes and the lasers and filters used in this confocal system.
For help in understanding how to choose the proper zoom factor, go to the optical resolution/zoom discussion. For help in understanding resolution in the z-axis, go to the page that discusses depth of field.
Life Sciences South Building, Room 444
1007 E. Lowell Street
Tucson, AZ 85721-0106
Phone:
520-621-7560
Fax: 520-621-3709
Email: MCB front office