Intracellular determination of elements in mammalian cultured cells by total reflection X-ray fluorescence spectrometry

TXRF (total reflection X-ray fluorescence spectrometry), a rapid and reliab le multi-element method, was applied to quantify intracellular concentratio ns of trace and non-trace elements in different mammalian cell lines. The c oordinated variations in their intracellular contents were determined when the extracellular concentration of one of them was modified. The results in dicate that TXRF permits the detection of total trace metal contents using a minimum amount of cells [(1-2) x 10(6)), while (4-6) x 10(6) cells were s ufficient to determine their cytosol/pellet distribution. In the six cell l ines analyzed, the order of relative abundance for trace and non-trace elem ents was Cu < Fe < Zn and Ca < S, respectively. One of the cell lines was e xposed to increased amounts of extracellular copper (0.44 to 100 mu mol l(- 1)), and then transferred to a culture medium containing a minimum concentr ation of the metal (0.44 mu mol l(-1)). Under this condition, it was possib le to measure both the rise (28-fold) in the intracellular copper content a nd its recovery to a base concentration. In addition, intracellular levels of other elements were measured, indicating that Fe and Ca showed the highe st increase (1.7- and 2.2-fold respectively). On the other hand, the Zn/Fe ratio was hardly affected, suggesting that this ratio can be used as an int ernal marker of metabolic integrity in cells exposed to different extracell ular copper concentrations. These results indicated that TXRF is an adequat e method to measure simultaneously the intracellular concentration of diffe rent elements and permits the evaluation of their variations in response to modified concentrations in the extracellular medium of one element.