An ultrasensitive method for the measurement of human leukocyte calcium: lymphocytes

Studies of the transport and distribution of calcium in leukocytes have been severely hampered by the inability to measure accurately and reproducibly the concentration of calcium in small numbers of cells. We have applied a recent development in analytical chemistry, the graphite furnace atomic absorption spectrophotometer, to this problem. The calcium content of human blood lymphocytes was determined by both graphite furnace and conventional flame atomic absorption spectrophotometry. The linearity, sensitivity and detection limits of the two techniques were compared. For measurement of calcium, the graphite furnace sensitivity was 55 times higher in aqueous samples and 60 times higher in cell samples than the flame technique. The detection limit of the graphite furnace was 800 times lower in aqueous samples and 1500 times lower in cell samples. The enhanced sensitivity of this technique allowed us to prepare samples with 20 times fewer blood cells. We have employed this graphite furnace technique to measure lymphocyte calcium content and its relationship to the calcium concentration and proportion of serum in the suspending medium. In the absence of serum, the lymphocyte calcium content approximately doubled as the medium calcium concentration was increased from 1 mumol/l to 0.5 mmol/l. At medium calcium concentrations of 0.5 mmol/l and above, the lymphocyte calcium content was 1.0 mmol/l cells. In medium adjusted to 2 mmol/l calcium, the lymphocyte calcium content approximately doubled as the medium serum concentration was increased from 0 to 2%. At medium serum concentrations of 2% and above, lymphocyte calcium content was 2 mmol/l cells. The exchangeable cell calcium, measured with 45Ca in the same samples, did not increase as serum was added to the medium.