Evaluation of toluidine blue for measuring erythrocyte membrane loss during in vivo ageing.

A simple method using a cationic dye, toluidine blue (TB), to quantify changes of red cell membrane area has been developed and tested for its validity. After incubating a glucose-depleted red cell suspension with a fixed quantity of TB at 37 degrees C for 10 min, the remaining TB was measured spectrophotometrically at 640 nm. Using this technique, we were able to show differences in TB uptake by populations of young and old red cells. The exact mechanism for TB uptake by the red cells is not clear. Treatment of the cells with bromelin, papain and trypsin reduced the uptake of TB, but neuraminidase and ficin had little or no effect. No inhibition of TB removal by red cells was observed using heparin, D-glucose, glucuronic acid, or N-acetylglucosamine.