Re: Paul's Tutorial on FL1, FL2, etc.

From: Emmanuel Gustin (Emmanuel.Gustin@skynet.be)
Date: Fri Dec 21 2001 - 18:20:22 EST


> Center of emissions wavelength: Incomplete (it has been already
acknowledged
> that band-width is important too) and entirely unnecessary on instruments
in
> which the optics are standard and specified by the manufacturer.

Only if you do all your work on a single instrument. But
in my experience people often look at the same cells with
other instruments, including fluorescence microscopes or
even readers, and then it is quite important to know what
the wavelength range is. It can be very difficult to make
any sense of quantitative results without that knowledge
--- often it is complicated enough even with it.

For the same reason, compensation settings should always
be documented and published. They may vary, yes, but only
within some margin; and you can always give an estimate
of this margin. If you don't include the compensation settings,
you are basically plotting an arbitrary quantity.

> our lab, and in all others with unmodified Coulter XLs cytometers,
> FL1 can be used perfectly intelligibly in an axis label as shorthand
> for: [...]

Within your lab, that can very well be an acceptable practice.
But "FL1 on Coulter XL" is of course only informative for people
who know the instrument. In publications I definitely want
to see a specification of the excitation wavelength and of
the central wavelength AND bandwidth of the emission filter.
The specifications of the dichroic optics might be omitted,
assuming that the optics designer has done his work well.
(And when you specificy which fluorochromes you use,
you should give a reference to a description of their spectral
properties.)

> certainly behooves me to detail this information in the materials and
> methods, but PLEASE, not in the axis label (see Chart Junk, below).

I would agree that this doesn't belong in the axis label.
It can be detailed in the text, or if you use several different
settings, in the figure caption. I would argue in favour
of putting it in the figure caption, for clarity.

Emmanuel Gustin



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