Single-Cell ICP-MS Analysis of Large Human Cells: Addressing Transport Efficiency and Size-Dependent Bias

Single-cell inductively coupled plasma-mass spectrometry (SC-ICP-MS) is a powerful tool for quantifying elemental contents in individual cells, providing insights into the levels of endogenous and exogenous elements. In SC-ICP-MS, cells are introduced into the plasma as highly diluted suspensions, generating transient events recorded in time-resolved mode. However, delivering cells intact into the plasma remains a challenge. The transport efficiency (TE) is never 100%, with cell size as a key limiting factor – an inverse relationship between the two has been reported. Although the use of specialized single-cell sample introduction glassware and chemical fixation improve cell integrity, these efforts remain insufficient for large human cells. Heated spray chambers have been explored, but while promising, their use remains limited. In Mass Cytometry, however, a heating block is integral to spray chamber, enabling TEs of 30–40% for various cell types.

Building on this strategy, this work explores the use of a custom-built heated spray chamber to enhance TE in SC-ICP-MS. In this case, a time-of-flight MS was used to allow simultaneous multi-element detection. The heated spray chamber significantly improved TE across various human cell lines (red blood cells, Raji cells, and A549 cells) when operated at temperatures above 100 °C. Most notably, the TE for A549 cells ( >20 µm) increased from 0.3% to 22.5%, enabling their analysis. Despite these improvements, TEs remain below 100%, which can indicate the potential size-dependent bias analysis within the same cell type population, thus potentially affecting the accuracy of single-cell measurements.

To investigate size-dependent sampling bias, laser ablation (LA)-ICP-TOF-MS is explored as an alternative, as it achieves 100% TE by sampling all cells regardless of size or morphology. However, LA sampling has lower throughput than SC-ICP-MS (hundreds vs. thousands of cells per experiment). To address this limitation, various cell deposition strategies—such as smearing, cytospinning, and single-cell array spotters—will be evaluated to enhance measurement efficiency. In parallel, membrane staining and tagging approaches (e.g., using wheat germ agglutinin or OsO₄) will be assessed as indicators of cell volume or size. These efforts will support a comprehensive comparison between LA-ICP-MS and SC-ICP-MS, providing insights into size-dependent sampling bias.