Immune Profiling and Multiplexed Label‐Free Detection of 2D MXenes by Mass Cytometry and High‐Dimensional Imaging

Abstract

There is a critical unmet need to detect and image 2D materials within single cells and tissues while surveying a high degree of information from single cells. Here, a versatile multiplexed label‐free single‐cell detection strategy is proposed based on single‐cell mass cytometry by time‐of‐flight (CyTOF) and ion‐beam imaging by time‐of‐flight (MIBI‐TOF). This strategy, “Label‐free sINgle‐cell tracKing of 2D matErials by mass cytometry and MIBI‐TOF Design” (LINKED), enables nanomaterial detection and simultaneous measurement of multiple cell and tissue features. As a proof of concept, a set of 2D materials, transition metal carbides, nitrides, and carbonitrides (MXenes), is selected to ensure mass detection within the cytometry range while avoiding overlap with more than 70 currently available tags, each able to survey multiple biological parameters. First, their detection and quantification in 15 primary human immune cell subpopulations are demonstrated. Together with the detection, mass cytometry is used to capture several biological aspects of MXenes, such as their biocompatibility and cytokine production after their uptake. Through enzymatic labeling, MXenes’ mediation of cell–cell interactions is simultaneously evaluated. In vivo biodistribution experiments using a mixture of MXenes in mice confirm the versatility of the detection strategy and reveal MXene accumulation in the liver, blood, spleen, lungs, and relative immune cell subtypes. Finally, MIBI‐TOF is applied to detect MXenes in different organs revealing their spatial distribution. The label‐free detection of 2D materials by mass cytometry at the single‐cell level, on multiple cell subpopulations and in multiple organs simultaneously, will enable exciting new opportunities in biomedicine.

Environmental and Health Impacts of Graphene and Other Two-Dimensional Materials: A Graphene Flagship Perspective

“…In particular, DCs showed the most prominent binding to these MXenes. 264 To sum up the literature to date, it remains challenging to conclusively determine the specific immune impact of 2D materials even within their individual subclasses due to minute differences in their manufacturing process, structure, functionalization or dispersibility profile, which could lead to various possibilities of cell interaction and downstream response. Macrophages remain the most popular immune cell of investigation due to their role as a key phagocytic cell of the innate immune system, as well as ease of culture.…”

“…Among the tested materials, Nb 4 C 3 displayed the strongest signal and extensive cellular interaction could be detected. In particular, DCs showed the most prominent binding to these MXenes …”

Latest advances on MXenes in biomedical research and health care

“… 21 , 48 Recently, high-dimensional immune and functional profiling of Nb 4 C 3 , Mo 2 Ti 2 C 3 , and Ta 4 C 3 T x has revealed that these forms of MXenes do not affect immune cell functionality and have excellent biocompatibility. 65 Furthermore, these MXenes possess intrinsic immunomodulatory properties that facilitate engineering the immune and inflammatory response of different cells and tissues (Figure 3 e). 60 , 61 For example, treating activated cocultures of human umbilical vein endothelial cells (HUVECs) and interferon-gamma (IFN-γ) expressing T-lymphocytes with Ta 4 C 3 T x QDs significantly reduced the percentage of IFN-γ + T-lymphocytes, indicating successful immunomodulation.…”

“…9 Other forms of MXenes, such as Nb 4 C 3 , Mo 2 Ti 2 C 3 , and Ta 4 C 3 T x are gaining traction for immunomodulation due to their excellent biocompatibility and intrinsic immunomodulatory properties. 65 …”

“…Pushing MXene-based biomedical technologies through the translational pipeline for eventual human use will necessitate a thorough evaluation of the biosafety, toxicity, and long-term stability of MXenes. Although detailed investigations of the biosafety of Nb 4 C 3 , Mo 2 Ti 2 C 3 , and Ta 4 C 3 were recently demonstrated, 65 similar analysis needs to be performed for other MXenes and in larger animal models over chronic time scales, in relevant use-case and form factor scenarios, and according to established standards. 74 Implantable bioelectronic devices and chronic biosensors will require strategies for minimizing the host immune response and to extend the material lifetime to the required time scales.…”