Single-particle (sp) mode analysis has been established in the ICP-MS field for the analysis of particle suspensions. This type of analysis uses a very short integration time of the mass spectrometer, usually less than 5 milliseconds, and dilutes the suspension (e.g. of nanoparticles) so that only a single particle is detected in this integration window. This approach forms the basis for a new trend that can currently be observed, in which particles are also measured in addition to biological and medical cells, but in this case with a diameter of > 5 µm. This technique is called single-cell analysis. Unlike with SP-ICP-MS, however, with this method it is recommended to use a syringe pump rather than a conventional peristaltic pump for sample transport, as cells can become crushed between the tube walls of the peristaltic pump, which can falsify the results of such “single-cell analyses”. An alternative sample feed method using a syringe pump is therefore recommended.
Syringe pumps are already used in many laboratories, especially when it comes to dosing or continuously transporting nanoliter or microliter volumes. Such applications require a particularly high level of precision, which is usually reflected in the purchase price of the corresponding technology. Delivery rates of up to 1,000 µL per minute, as required for ICP-AES or ICP-MS, are therefore more of an exception. Spetec has therefore developed a syringe pump that meets the special requirements of atomic spectroscopy and offers an excellent price-performance ratio. This pump was used in a study by Gunda Köllensperger’s working group at the University of Vienna on the uptake and kinetics of new cytostatics in the treatment of cancer cells. The method is based on a time-resolved single-cell analysis in which a time-of-flight mass spectrometer (icpTOF 2R; TOFWERK AG, Thun, Switzerland) is used instead of a conventional quadrupole-based mass spectrometer. ICP-TOF-MS offers the advantage of providing a true multi-element analysis of the individual cells, since sequentially measuring quadrupole or sector field instruments only provide the time-resolved analysis of a single isotope. For this experiment, yeast cells (pichia pastoris) were incubated with cisplatin (5 µM, 37°C, 24h). The cells were then washed and filled into the syringe of the syringe pump. The flow rate of 10 µL per minute was adapted to the nebulizer used (Parallel Path PFA 260, AHF Analysentechnik, Tübingen). The ICP-TOF-MS simultaneously records the data of all isotopes in the mass range m/Z 2 – 257 and with appropriate calibration the measured intensity can be directly converted into femtograms per single cell. This study was able to show that it is possible to determine the presence of metals in the low femtogram range in individual cells. The masses determined for the elements in individual cells correlated closely with the decomposed average values of the results of an ICP-MS analysis performed on a large number of cells. However, when the results of such multi-cell analyses are broken down, important information is lost, namely the heterogeneity of the cell system, since the distribution is not Gaussian, which can be seen from the fact that individual cells can absorb disproportionately large amounts of cisplatin. However, if one compares this cisplatin distribution with that of other essential elements (Fe, Cu, Zn), one can see that this special distribution is due to the biological variability of the cells. Overall, this study shows that careful sample supply using a syringe pump significantly simplifies single-cell analysis using ICP-MS and enables detection limits for Cu and Zn in the lower attogram range and for P and Fe in the lower femtogram range. With the method presented here, it will now be possible to examine other medically relevant cell systems for their interaction with new, metal-containing cytostatics. Of course, such investigations are also possible with conventional ICP-MS devices. However, it is necessary to measure the individual isotopes of the metals of interest one after the other, which significantly increases the analysis time. This method is not only applicable in pharmaceutical research, but also in toxicological studies to clarify the transport of nanoparticles or toxic heavy metals in cells.
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