Selection of Fluorochromes
Fluorochrome selection should be carefully carried out using the Guides below.
The Dyes with the highest Staining Index should be used with the weakest antigen expression, giving you the best chance of detecting the different antigens from autofluorescence. You should never just add another Fluorochrome, without assessing the expression levels as this will lead to the weak fluorochromes on the weakest antigens.
The spectral overlap of fluorochomes should also be taken into consideration when planning an experiment, as poor combinations may affect the clarity of the results.
When performing compensation if possible use the software autocompensation as this gives scientifically correct results, but in combination with your single colour controls you will need ‘all but one’ controls to specify the negative/ positive boundary. Further information can be found on the BD , Biolegend web sites