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Free radical generation plays a key role in killing bacteria by antibiotics. However, radicals are short-lived and reactive, and thus difficult to detect for the state of the art. Here we use a technique which allows optical nanoscale magnetic resonance imaging (MRI) to detect radical generation on the scale of single bacteria. We demonstrate that the radical generation in Staphylococcus aureus increases in the presence of UV irradiation as well as vancomycin and is dependent on the antibiotic's dose. With a method based on ensembles of nitrogen vacancy (NV) centers in diamond, we were able to follow the radical formation near individual bacteria over the whole duration of the experiment to reveal the dynamics of radical generation. Using this new approach, we observed free radical concentrations within nanoscale voxels around the diamond particles and determined its exact timing depending on the antibiotic dose. Since changes in the response to antibiotics emerge in only a few bacteria of the entire population, such a single-cell approach can prove highly valuable for research into drug resistance.

Free radicals play a crucial role in numerous diseases. Currently, very little is known about where and when radicals are build and how they work. Their short lifetime and reactivity make measuring them challenging. There are currently several methods available to detect free radicals inside cells; however none of them can determine the location and detect the radicals at high time resolution. Nanodiamonds have superior physical and chemical properties and are excellently biocompatible. The NV centers make it possible for nanodiamonds to function as biosensors to detect free radicals. NV centers can convert magnetic noise into an optical signal by changing their brightness according to the magnetic surroundings and allowing radical detection in real-time.

Diamond magnetometry makes use of fluorescent defects in diamonds to convert magnetic resonance signals into fluorescence. Because optical photons can be detected much more sensitively, this technique currently holds several sensitivity world records for room temperature magnetic measurements. It is orders of magnitude more sensitive than conventional magnetic resonance imaging (MRI) for detecting magnetic resonances. Here, the use of diamond magnetometry to detect free radical production in single living cells with nanometer resolution is experimentally demonstrated. This measuring system is first optimized and calibrated with chemicals at known concentrations. These measurements serve as benchmarks for future experiments. While conventional MRI typically has millimeter resolution, measurements are performed on individual cells to detect nitric oxide signaling at the nanoscale, within 10–20 nm from the internalized particles localized with a diffraction limited optical resolution. This level of detail is inaccessible to the state-of-the-art techniques. Nitric oxide is detected and the dynamics of its production and inhibition in the intra- and extracellular environment are followed.

We demonstrated nanoscale MRI of metabolism in single isolated mitochondria and within living cells using NV centers in diamond., 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.

Diamond magnetometry is a quantum sensing method involving detection of magnetic resonances with nanoscale resolution. For instance, T1 relaxation measurements, inspired by equivalent concepts in magnetic resonance imaging (MRI), provide a signal that is equivalent to T1 in conventional MRI but in a nanoscale environment. We use nanodiamonds (between 40 and 120 nm) containing ensembles of specific defects called nitrogen vacancy (NV) centers. To perform a T1 relaxation measurement, we pump the NV center in the ground state (using a laser at 532 nm) and observe how long the NV center can remain in this state. Here, we use this method to provide real-time measurements of free radicals when they are generated in a chemical reaction. Specifically, we focus on the photolysis of H2O2 as well as the so-called Haber-Weiss reaction. Both of these processes are important reactions in biological environments. Unlike other fluorescent probes, diamonds are able to determine spin noise from different species in real time. We also investigate different diamond probes and their ability to sense gadolinium spin labels. Although this study was performed in a clean environment, we take into account the effects of salts and proteins that are present in a biological environment. We conduct our experiments with nanodiamonds, which are compatible with intracellular measurements. We perform measurements between 0 and 10(8) nM, and we are able to reach detection limits down to the nanomolar range and typically find T1 times of a few 100 mu s. This is an important step toward label-free nano-MRI signal quantification in biological environments.