🚀 go-pugleaf

We have an MRC-600 with a K-Ar laser source and in anticipation of doing
sequential triple labelling  bought a trichroic block which goes in the K1
position. In conjunction we have two purpose built "paddle stick" filters
designed with filter (605/32 and 680/32) windows which are inserted in port
S5 in between the K2 block and PMT1. This allows one to discriminate
between the Texas Red emission and Cy5 emission from an appropriately
labelled and excited piece of tissue. It thus allows me to look at the
bleed thru from an FITC labelled specimen. Standard bleed thru with no
paddle stick is  reduced with the 605/32 window and negligible if the
680/32 paddle stick is in place (I have to turn gain and PMT aperture very
high and wide to see any specimen at all). I suggest to clients to use FITC
and Cy5 wherever practical as their double label if lack of cross talk is
crucial (hence my desire to have a good red line!). Other than that I
suggest the client titrate their FITC histochemistry to give a clearly
discernible but not overpowering signal or to use the FITC probe against
the smaller better defined moeity in the specimen. Hope this is of some
use.
                Cheers,   Colin




The Hielander
Colin Macqueen      C.L.Macqueen@vthrc.uq.edu.au
Confocal Facility   The University of Queensland  Fax +61-7-365-4522