Why is my FAM labeled oligo not as bright as it was 6 months ago?
Under proper conditions, fluorescein-based dyes (e.g., 6-FAM) have excellent signal intensity for a range of applications. However, they are susceptible to a relatively high rate of photobleaching and are sensitive to acidic environments, which can diminish the signal over time. The signal will rapidly degrade as the dye becomes protonated, therefore FAM-labeled oligonucleotides should remain in solutions with 7.2 pH or higher to limit degradation. To minimize photobleaching of fluorescein-labeled oligos, we recommend using a maximally sensitive detection system with the lowest possible intensity excitation used in tandem with broad bandpass filters. Fluorophores such as Alex Fluor 488, ATTO 488, BODIPY FL, Oregon Green 488, Oregon Green 514 and Rhodamine Green dyes are more photostable than fluorescein and less pH sensitive in the physiological pH range, making them good alternatives for some applications. More information on dyes and their chemical stability can be found at the Molecular Probes website.