Quantum monitoring of cellular metabolic activities in single mitochondria.

Free radicals play a vital role in all kinds of biological processes including immune responses. However, free radicals have short lifetimes and are highly reactive, making them difficult to measure using current methods. Here, we demonstrate that relaxometry measurement, or T1, inherited from the field of diamond magnetometry can be used to detect free radicals in living cells with subcellular resolution. This quantum sensing technique is based on defects in diamond, which convert a magnetic signal into an optical signal, allowing nanoscale magnetic resonance measurements. We functionalized fluorescent nanodiamonds (FNDs) to target single mitochondria within macrophage cells to detect the metabolic activity. In addition, we performed measurements on single isolated mitochondria. We were able to detect free radicals generated by individual mitochondria in either living cells or isolated mitochondria after stimulation or inhibition.

Nanodiamonds as multi-purpose labels for microscopy.

Nanodiamonds containing fluorescent nitrogen-vacancy centers are increasingly attracting interest for use as a probe in biological microscopy. This interest stems from (i) strong resistance to photobleaching allowing prolonged fluorescence observation times; (ii) the possibility to excite fluorescence using a focused electron beam (cathodoluminescence; CL) for high-resolution localization; and (iii) the potential use for nanoscale sensing. For all these schemes, the development of versatile molecular labeling using relatively small diamonds is essential. Here, we show the direct targeting of a biological molecule with nanodiamonds as small as 70 nm using a streptavidin conjugation and standard antibody labelling approach. We also show internalization of 40 nm sized nanodiamonds. The fluorescence from the nanodiamonds survives osmium-fixation and plastic embedding making them suited for correlative light and electron microscopy. We show that CL can be observed from epon-embedded nanodiamonds, while surface-exposed nanoparticles also stand out in secondary electron (SE) signal due to the exceptionally high diamond SE yield. Finally, we demonstrate the magnetic read-out using fluorescence from diamonds prior to embedding. Thus, our results firmly establish nanodiamonds containing nitrogen-vacancy centers as unique, versatile probes for combining and correlating different types of microscopy, from fluorescence imaging and magnetometry to ultrastructural investigation using electron microscopy.