Conjugates of AMCA absorb light maximally at 350 nm and fluoresce maximally at 450 nm (Table 1 and Figure 2). For fluorescence microscopy, AMCA can be excit-ed with a mercury lamp and observed using a UV filter set. Since blue fluorescence is not well detected by the human eye, AMCA-conjugated secondary antibodies should be used with the most abundant antigens in multiple-labeling experiments. Ways of improving the visibility of AMCA include dark adapting the eyes, using fluorite instead of glass objectives, avoiding mounting media that absorb UV light (such as plastic-based media), and capturing photographic images. AMCA also fades as rapidly as fluorescein; and therefore, it should be used with an anti-fading agent. 

For flow cytometry, AMCA can be excited with a mercury lamp, or with a water-cooled argon ion laser which emits some lines in the UV region of the spectrum.

Table 1: Approximate* peak wavelengths of absorption and emission for different fluorophore-conjugated, affinity-purified antibodies.

AMCA has been used mostly for multiple labeling since there is minimal fluorescence overlap with fluorescein and little or no overlap with longer wavelength-emitting fluorophores. Applications for multiple labeling with this probe include both immunofluorescence microscopy and flow cytometry. AMCA is not suggested for single labeling because of its relatively weak signal.