In order to compare the recovery ef

In order to compare the recovery efficiencies, we obtainedintracellular extracts of MCF-7 cells using both the newcell quenching method reported here and the conventionalmethod. Shown in Fig. 1a is the conventional samplepreparation method for adherent cells (Beloueche-Babariet al. 2006; Lane and Fan 2007; Yang et al. 2007). Afterremoving the medium and rinsing with PBS, trypsin is usedto detach the cells from their growth surface. The detachedcells are then washed with PBS and centrifuged 2 or 3times to minimize the carryover of culture medium beforea quench solution (methanol is typically used) is added tothe cells.This contrasts with the new method shown in Fig. 1b,which involves direct quenching of adherent cells. Trypsin,which invariably changes the extracellular environment ofthe cells (as shown visually in Fig. 1c), is not used. Cellsare quickly rinsed with ice-cold PBS twice after the mediumis removed, and quenched using methanol. It requires20 s to process cells using the new direct cell quenching