This is consistent with our findings on an MRC-600.

________________________________________________________________________________


Paul Goodwin
Image Analysis Lab
FHCRC, Seattle, WA

On Mon, 20 Jan 1997, Stephen T. Haley, II wrote:

> Hello List Members,
>      I have already sent this to the capable people at Bio-Rad but
> thought I may post it to the list for some feedback.  Thanks in
> advance for any ideas.
>
>      I am trying to do some "relative" quantitative immunoflourescence
>
> on FITC labelled tissue sections using a Bio-Rad MRC-1000.
> I was happily collecting data.
> The levels of the surface markers I am trying to measure went up or
> down as I expected. The trouble arose when I tried to check the
> validity of my measurements with some InSpeck beads from Molecular
> Probes.  The beads are calibrated to several relative intensities. To
> make a long story short I found out that the system is not linear. I
> am working in the lower portion of the 0-255 scale. The laser settings
> Iam using are as follows in Photon counting mode: Iris=3.0,
> Gain=1250,Multiplier=6, and the Laser set to 0.3, 1% or 3%, depending
> on the relative brightness of the bead. I have played around with the
> recommended zoom settings (from Pawley's handbook) to satisfy the
> Nyquist criterion but it does not seem to help.
>
> Thanks in Advance for your suggestions,
>
>
> Stephen T. Haley
> Graduate Student
> University of South Carolina School of Medicine
> Department of Microbiology and Immunology
>