Delicate and crucial pH dependence in permeabilization of cultured mammalian cells by vortex-stirring with a high molecular weight polyacrylic acid.

We previously reported that cultured mammalian cells are effectively permeabilized by vortex-stirring the cells with a high molecular weight polyacrylic acid. The present study revealed that the efficiency of permeabilization of a non-permeant dye, Lucifer Yellow (LY), was sensitively affected by the pH of the medium. When the cells are vortex-stirred in RPMI 1640 culture medium, the pH of the medium should be adjusted with HEPES and NaOH, instead of NaHCO3, to a pH higher than 7.6 for successful permeabilization; the higher the pH, the better the result obtained. Internalization of LY was near the background level at a pH below 7.4. When phosphate-buffered saline was substituted for RPMI 1640 medium, the optimal pH range was slightly shifted to a more acidic region. This requirement for the pH of the medium is indispensable as a supplement to the standard permeabilization procedure tentatively proposed in the preceding report.