ABSTRACT
3D super-resolution microscopy with nanometric resolution is a key to fully complement ultrastructural techniques with fluorescence imaging. Here, we achieve 3D super-resolution by combining the 2D localization of pMINFLUX with the axial information of graphene energy transfer (GET) and the single-molecule switching by DNA-PAINT. We demonstrate <2 nm localization precision in all 3 dimension with axial precision reaching below 0.3 nm. In 3D DNA-PAINT measurements, structural features, i.e., individual docking strands at distances of 3 nm, are directly resolved on DNA origami structures. pMINFLUX and GET represent a particular synergetic combination for super-resolution imaging near the surface such as for cell adhesion and membrane complexes as the information of each photon is used for both 2D and axial localization information. Furthermore, we introduce local PAINT (L-PAINT), in which DNA-PAINT imager strands are equipped with an additional binding sequence for local upconcentration improving signal-to-background ratio and imaging speed of local clusters. L-PAINT is demonstrated by imaging a triangular structure with 6 nm side lengths within seconds.
ABSTRACT
DNA nanotechnology offers new biosensing approaches by templating different sensor and transducer components. Here, we combine DNA origami nanoantennas with label-free antibody detection by incorporating a nanoswitch in the plasmonic hotspot of the nanoantenna. The nanoswitch contains two antigens that are displaced by antibody binding, thereby eliciting a fluorescent signal. Single-antibody detection is demonstrated with a DNA origami integrated anti-digoxigenin antibody nanoswitch. In combination with the nanoantenna, the signal generated by the antibody is additionally amplified. This allows the detection of single antibodies on a portable smartphone microscope. Overall, fluorescence-enhanced antibody detection in DNA origami nanoantennas shows that fluorescence-enhanced biosensing can be expanded beyond the scope of the nucleic acids realm.
ABSTRACT
We introduce p-MINFLUX, a new implementation of the highly photon-efficient single-molecule localization method with a simplified experimental setup and additional fluorescence lifetime information. In contrast to the original MINFLUX implementation, p-MINFLUX uses interleaved laser pulses to deliver the doughnut-shaped excitation foci at a maximum repetition rate. Using both static and dynamic DNA origami model systems, we demonstrate the performance of p-MINFLUX for single-molecule localization nanoscopy and tracking, respectively. p-MINFLUX delivers 1-2 nm localization precision with 2000-1000 photon counts. In addition, p-MINFLUX gives access to the fluorescence lifetime enabling multiplexing and super-resolved lifetime imaging. p-MINFLUX should help to unlock the full potential of innovative single-molecule localization schemes.
Subject(s)
Nanotechnology , Photons , DNA , Lasers , Microscopy, FluorescenceABSTRACT
AIMS AND OBJECTIVES: To evaluate the effect of a stop smoking clinic on the quit rates of patients admitted to an acute in-patient unit. BACKGROUND: The relationship between poor physical health and severe mental illness is well established. High rates of smoking appear to play an important causal role in the excess morbidity and mortality in this population. Stop smoking interventions for the general population are clinically effective and cost-effective. There is a small but promising evidence base for effective interventions to help people with a mental illness who wish to stop smoking but these have mostly been tested with community patients rather than acute in-patients. METHODS: A service evaluation of a drop-in stop smoking clinic on an acute mental health in-patient unit was conducted. Patients' smoking status was measured at baseline and four weeks after their quit date using patient self-report and an expired breath carbon monoxide reading. RESULTS: Over a six-month evaluation period, 46 patients set a quit date and 13 (28·3%) were abstinent at the four-week follow-up stage, verified by a carbon monoxide reading (χ(2) =33, df=1, sig p<0·0001). CONCLUSIONS: This small-scale evaluation has shown a drop-in stop smoking intervention to be feasible, acceptable and associated with positive outcomes; further research with larger, more representative samples is required. RELEVANCE TO CLINICAL PRACTICE: Enforcing smoke-free legislation is a contentious issue on mental health in-patient units, and there is a paucity of research to guide nursing practice in this area. An admission period in a smoke-free environment provides a crucial opportunity to offer smoking cessation treatment. With appropriate resources, expertise and support, it appears possible to apply smoking cessation interventions that are successful within the general population to mental health patients during an acute admission.
