Is it safe to fix stained samples?

In a perfect world, everything would go smoothly, we’d only work with clean samples and they could be harvested, stained and acquired all in a day’s work. Unfortunately, science is never predictable. Researchers need to be able to fix their samples due to the presence of pathogens or to simply save the samples for analysis at a later time. Many researchersare reluctant to fix samples (unless they are required to)because they worry about fluorochrome stability, (especially tandems) and havebeen told that tandem dyes fall apart after fixation. THIS IS A MYTH. Tandem dyes are two fluorochromes covalently linked, making them impossible to fall apart,however when handled improperly, they can lose FRET (Fluorescence ResonanceEnergy Transfer) efficiency which results in a need for higher compensation.Several factors can affect FRET efficiency.  Recommended precautions are listed to help minimize FRET loss.

Affected By:   Resolution:

light exposure protect samples from direct light

oxygen exposure consider degassing flow buffers

poor quality fixatives use eBioscience Fixation buffers

We understand that fixation may be a requirement with certain samples or whenimmediate analysis is simply not an option. If samples stained with antibodiesconjugated to organic fluorochromes must be stored, we recommend completionof the staining protocol and fixation of samples with IC Fixation Buffer (cat. 008222)(100 µL sample with 100 µL IC Fixation Buffer) or 1-step Fix/Lyse Solution(cat. 00-5333) (100 µL sample with 2 mL 1-Step Fix/Lyse Solution). Cells can bestored in these buffers for up to 3 days in the dark at 4°C with minimal impact onbrightness or FRET efficiency/compensation when using the IC Fixation (cat. 008222)or 1-step Fix/Lyse Solution (cat. 00-5333).