Label-free metabolic and structural profiling of dynamic biological samples using multimodal optical microscopy with...

Area of Science:

• Biomedical Optics
• Microscopy
• Label-Free Imaging

Background:

• Label-free optical microscopy offers noninvasive biological imaging but faces limitations in specificity, speed, and signal quality.
• Existing techniques struggle with accurate quantification due to signal noise and slow processing.

Purpose of the Study:

• To introduce FOCALS (Fast Optical Coherence, Autofluorescence Lifetime imaging, and Second harmonic generation) microscopy, a novel multimodal label-free imaging technique.
• To overcome the limitations of traditional label-free microscopy by enhancing specificity, speed, and quantitative accuracy.

Main Methods:

• Simultaneous acquisition of NAD(P)H fluorescence lifetime, second harmonic generation (SHG), and polarization-sensitive optical coherence microscopy (OCM) images.
• Implementation of fast analog detection and real-time GPU processing for longitudinal imaging.
• Integration of sensorless adaptive optics for aberration correction in thick tissues.

Main Results:

• FOCALS microscopy provides quantitative metabolic and morphological profiles across various sample types (in vitro, ex vivo, in vivo).
• Imaging speed is increased 4-40x with real-time processing, maintaining signal integrity for quantitative nonlinear microscopy.
• Adaptive optics significantly improves imaging quality in thick tissues, enhancing quantitative fluorescence imaging.

Conclusions:

• FOCALS microscopy overcomes key limitations of traditional label-free imaging, offering improved specificity and quantitative analysis.
• The multimodal approach and advanced processing enable faster, more accurate, and versatile biological imaging.
• This technique facilitates longitudinal studies of biological dynamics with enhanced resolution and reduced artifacts.