Trypan Blue is the stain most commonly used to distinguish viable from nonviable cells. The reactivity of trypan blue is based on the fact that the chromophore is negatively charged and does not interact with the cell unless the membrane is damaged. Therefore, all the cells which exclude the dye are viable, while nonviable cells absorb the dye and appear blue. Cells should be in suspension as single cells in buffered saline before counting. Trypan Blue has a higher affinity for serum protein than for cellular proteins, so suspending cells in medium containing serum will generate a dark background.