Re: dWater as sheath plus ARIA question

Alice Givan wrote:

>To Howard et al,
>As Howard said, you cannot use distilled water in the sheath stream 
>if you are sorting as
>distilled water will not take the charge needed for sorting.
>
>But, even for non-sorting applications,  I have always thought 
>(although have not tested)
>that using distilled water as sheath fluid will raise	background 
>signals --- because
>light will bounce off the interface between the core stream and the 
>sheath fluid  (they
>no longer have the same refractive index).

Alice is right about light being scattered at the boundary of the 
core and sheath streams if the refractive indices are different, as 
is the case when a distilled water sheath is used for analysis of a 
sample in a saline solution. The background of *scatter* signals is 
then increased, although, at least in my experience, this becomes a 
problem only when one analyzes particles with very small scatter 
signals, such as bacteria. As was suggested in my earlier posting on 
this subject, I can usually get away with using a distilled water 
sheath even for analyses of bacteria. The Technicon/Bayer hematology 
instruments, which measure both scatter and absorption (highly 
sensitive to index mismatch), benefit from the use of index-matched 
sheaths, or at least the older ones did.

I suspect that any difference in fluorescence background due to 
core-sheath refractive index differences is small; I can get 1-2% CVs 
in fluorescence measurements of dyed beads or Hoechst dye-stained 
nuclei without matching refractive indices of core and sheath.

-Howard