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The Magic of Super-Resolution Microscopy: How to Break the Limits of Light?
In the scientific community, the advancement of microscopy technology is undoubtedly an important tool for revealing the mysteries of the microscopic world, among which super-resolution microscopy technology is particularly eye-catching. This series of technologies not only breaks through the diffraction limit of optical microscopes, but also demonstrates great potential in applications in biomedical research and molecular biology, providing us with a more comprehensive understanding of the internal structure and function of cells.
Super-resolution imaging techniques rely on the choice of either near-field (such as photon tunneling microscopy and near-field scanning optical microscopy) or far-field settings.
Super-resolution microscopy can be divided into two main categories: deterministic super-resolution technology and stochastic super-resolution technology. The former uses the nonlinear response of luminophores (fluorescent molecules) commonly used in biological microscopes to enhance resolution. Typical techniques include stimulated luminescence depletion (STED) and ground state depletion (GSD). The latter uses the temporal behavior of molecular light sources to enable similar fluorescent molecules to emit light separately, forming resolvable images. Such techniques include super-resolution optical wave imaging (SOFI) and single-molecule localization microscopy (SMLM). For example, PALM and STORM.
On October 8, 2014, Eric Büttig, Walter Molnar and Stefan Hell were awarded the Nobel Prize in Chemistry for "the development of super-resolved fluorescence microscopy", marking the first major breakthrough in the field of optical microscopy. Entering the nanoscale realm.
Theories for breaking the Abbe limit have been emerging since the 1970s. A 1978 research paper proposed the concept of using 4Pi microscopy, a laser scanning fluorescence microscope that achieves high resolution by focusing light sources from both sides. However, the research at that time did not pay enough attention to the improvement of axial resolution. In 1986, stimulated emission-based super-resolution optical microscopy was first patented.
Application of super-resolution technology
These super-resolution techniques not only provide new perspectives for microscopy, but also speed up the observation of biomolecules. Among them, the near-field optical random mapping (NORM) microscope obtains optical near-field information by observing the Brownian motion of nanoparticles in suspension. Its imaging process does not require special positioning equipment, which undoubtedly improves the efficiency of image acquisition.
Structured illumination microscopy (SIM) achieves enhanced spatial resolution by collecting frequency-spatial information outside the visible region, and has great potential for some medical diagnoses.
Mirroring these technological advances, structured illumination microscopy (SIM) has shown the potential to replace electron microscopy for certain medical diagnostics. For example, SIM is increasingly used in the study of kidney and blood diseases in medical diagnosis. In addition, spatially modulated illumination (SMI) further improves the accuracy of distance measurements, enabling molecular size measurements on the scale of tens of nanometers.
Application of biosensor technology in super analysis
In cell biology, biosensing technology is an important means to understand the activities of cellular components. These sensors usually consist of two parts: sensing and reporting, using fluorescence detection technology to quantify biological activities. The emergence of new fluorescent probes has greatly expanded the possibility of observing dynamic processes within cells.
REversible Saturable OpticaL Fluorescence Transitions (RESOLFT) microscopy not only enables the capture of more details in images, but also expands the concept of super-resolution, making it increasingly important in biomedical research.
With the continuous development of technology, deterministic methods such as STED and GSD have been gradually improved, providing new solutions. However, the practicality of these technologies is still challenged by the complexity of equipment and the risk of sample damage. Therefore, although super-resolution microscopy technology has extraordinary resolution capabilities, scientists still need to continue to explore its optimal application in various fields.
The integration and application of these technologies allow us to more intuitively understand cell machinery, structure and function, and ultimately inspire further biomedical research. How will future scientific discoveries expand our understanding of life? Woolen cloth?
