Electron probe X-ray microanalysis of cultured epithelial tumour cells with scanning electron microscopy.

Three methods have been used to prepare cultured cells for electron probe X-ray microanalysis (EPXMA): (1) analysis at the subcellular level of freeze-dried ultrathin cryosections with scanning transmission electron microscopy (STEM); (2) analysis at the cellular level of whole freeze-dried cells with STEM; and (3) analysis at the cellular level of whole freeze-dried cells with scanning electron microscopy (SEM) (for a review see: Wroblewski and Roomans, 1984; Wroblewski and Wroblewski, 1993; Warley, 1994). However, EPXMA of whole freeze-dried cells with SEM has some disadvantages. This method requires that cultured cells be adapted to growth on a thick substrate such as plastic or glass coverslips (Zierold and Schäfer, 1988), graphite discs (Abraham et al., 1985; Larsson et al., 1986) and microcarrier beads (Hall et al., 1992). In addition, a suitable washing procedure to remove the extracellular medium must be found because the deposition of culture medium on the cell surface after freezing and freeze-drying may interfere with X-ray spectra from the cell. Moreover, it is difficult to perform absolute quantitative analyses of elemental content, since the substrate may contribute to the background, decreasing the peak-to-background (P/B) ratio (Roomans, 1981). We present here a simple method to study the intracellular concentrations of elements in whole cultured epithelial tumour cells by EPXMA with scanning electron microscopy.