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UND 363 Aur/Rhodamin

UND 363 Mwave Auramine Rhoadmine Flourescence

QuestionAnswer
principle for Mwave Auramine Rhoadmine Flourescence demo myco tuberculosis or other A/F organisms
Principle dyes used are basic that floursece at short wavelengths. Using both dyes in combo will yield better results than alone (auramine O & Rhodamine B)
how do the bacteria gain the flourescence to be seen mycolic acid in the cell wall stains with the flourochromes in the dyes.
fix NBF preferred
micron 4-5 (will also need a flourescent microscope) use 40x hi-dry with exciter filter and colorless UV barrier filter
qc tissue with A/F organisms, must use Millipore filtered (.45 micron pore size) water, and a negative control from same day workload.
reagents in order auramineO-RhodamineB soln', acid alcohol - decolor/diff'r, eriochrome black T - counterstain
results A/F bacteria - red (yellow flourescence), background - black
why is this stain prone to false positives it is extremely sensitive and highly specific therefore it is difficult for inexperienced microscopists
if results are questionable with the aur/rho stain what can be used in addition carbol fuchsin can be used to restain for confirmation of the results (it is NOT possible to then go back to using Aur/rho after carbol fuchsin has been used)
if there are dead and dying organisms will they still stain yes, it will still stain dead and dying more so than the carbol-fuchsin method
why is it important to only use a small amount of rhodamine B? too large a quantity can quench flourescence (even in low conc.)
what can occur if rhodamine B concentration is reduced it will intensify the flourescens of mycobacteria but it will change the flourescence of the auramine o from yellow to orange-yellow and intensifies it.
what formalin soln's can't be used in this stain and why zine formalin. it will inhibit flourescence microscopy due to quenching.
Created by: mustangvxd
 

 



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