Keywords: confocal microscope, 3D imaging, in vivo observation, multiphoton microscope

Confocal microscopy filters out non focal plane signals through pinholes to achieve optical slicing and 3D reconstruction. Its core advantages include high resolution, high contrast, and tomographic capabilities, which are widely used in cell biology and neuroscience.

1、 The core principle and imaging mode of confocal microscope

Confocal microscope scans the sample point by point through laser, and the pinhole only allows the focal plane signal to pass through the detector. Its resolution is 2-3 times higher than that of a wide field microscope, making it suitable for tomographic imaging of thick samples. For example, in histological research, confocal microscopy can reconstruct cell migration pathways during embryonic development.

Multiphoton microscopy utilizes long wavelength lasers to excite fluorescent molecules, reducing light scattering and phototoxicity, and is suitable for imaging deep tissues in vivo. For example, in neuroscience, multiphoton microscopy can observe the calcium signals of neurons in the mouse cerebral cortex, revealing the neural mechanisms of learning and memory.

2、 Fluorescence Resonance Energy Transfer (FRET) Technology

FRET detects changes in intermolecular distance through energy transfer between donor and acceptor fluorescent molecules. For example, in signal transduction research, FRET can monitor the conformational changes of receptor proteins and reveal the mechanism of drug action.

3、 In vivo imaging and drug development

Confocal microscopy has important applications in live imaging. For example, in tumor research, confocal endoscopy can observe real-time angiogenesis and drug penetration in tumor tissue; In drug screening, high-throughput confocal platforms can simultaneously monitor the response of thousands of cells to drugs, accelerating the development of new drugs.

4、 Technological bottlenecks and future directions

The current challenges include phototoxicity, imaging depth, and data analysis complexity. Future trends include:

Adaptive optics technology: compensates for aberrations and improves the quality of deep tissue imaging.

Photogenetics combination: utilizing light controlled ion channels to achieve precise regulation and imaging of neural activity.

Miniature confocal system: Developing portable devices to achieve real-time clinical diagnosis.