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Pherastar - does anyone use one?

  • 12-12-2014 9:49am
    #1
    CramCycle
    Moderators, Sports Moderators Posts: 25,515 Mod ✭✭✭✭


    I am using a pherastar plate reader but the fluoresence readings I am getting are not as good as they are on other machines. I have read the manual on optimisation but it mentions using a "free tracer" to work out the optimal focal height and gain settings.

    My probe has a quencher attached so I presume throwing that in on its own isn't a "free tracer" (I don't even know what it is), do I just get a sample of the fluorophore unattached to anything and stick it in a well? I can't seem to figure it out.

    Has anyone here done it before, I used to use a Victor 2 which was easier as depending on the plate etc. it would just give you settings to throw in, the Pherastar seems to want to optimise itself but I am seemingly to stupid to do this.

    Any help appreciated.


Comments

  • #2
    cyberwolf77
    Moderators, Science, Health & Environment Moderators, Social & Fun Moderators Posts: 47,851 Mod ✭✭✭✭cyberwolf77


    I dug through the pdf of the manual online and it looks like the "free tracer" is something that you add to the assay plate before running it through. Please bear in mind on this topic I am a clever monkey with a hammer and a coconut.
    http://tools.lifetechnologies.com/content/sfs/manuals/PHERAstar_Predictor.pdf (pages 15-17)seem most relevant to your problem

    Hopefully this was of some assistance to you.


  • #3
    CramCycle
    Moderators, Sports Moderators Posts: 25,515 Mod ✭✭✭✭CramCycle


    I found the same, the best I can assume is that the positive control is a known PCR run with released fluorophore and the free tracer is just the fluorophore unbound to anything so there is no quenching.

    Guess I will just have to order in some unbound fluorophores, hooray for wasting money. Thanks for the help.


  • #4
    cyberwolf77
    Moderators, Science, Health & Environment Moderators, Social & Fun Moderators Posts: 47,851 Mod ✭✭✭✭cyberwolf77


    Best of luck to you, and please post back and let me know if it worked for you


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