SHG Microscopy for Collagen Imaging
Stain-Free Precision
SHG microscopy is a laser-based imaging technique that allows visualization of non-centrosymmetric structures like fibrillar collagen in tissues without the use of exogenous stains or dyes. This optical phenomenon occurs when two photons interact with a non-centrosymmetric structure and combine to generate a new photon with exactly twice the energy and frequency.
Revealing Hidden Structures - How SHG works
Next-generation Imaging
In SHG microscopy, tissues are exposed to femtosecond laser pulses. When these pulses interact with fibrillar collagen, the asymmetry in the structure causes the combining of two lower energy photons into a single higher energy photon. This emitted photon has exactly half the wavelength of the excitation laser. Special filters enable detecting these emitted photons to generate high contrast, high resolution images showing the distribution of fibrillar collagen.
As SHG signal generation is a physics phenomenon rather than one that depends on chemical dyes or stains, it avoids issues with staining variation, photobleaching and provides output images with intrinsic contrast in a highly consistent manner. The SHG intensity is also directly proportional to the quantity and organization of fibrillar collagen, allowing quantitative analysis.
Enabling Non-Destructive Collagen Monitoring
Continuous Fibrosis Profiling
Compared to traditional histological stains like Picosirius Red or Masson’s Trichrome, SHG provides intrinsic signals specific to fibrillar collagen structure. This eliminates variability due to stain penetration or protocol differences.
Since there is no photobleaching, repeated imaging is possible without signal loss. Furthermore, the non-destructive nature also preserves tissues for subsequent analyses. Quantification of SHG signals provides objective measures of fibrillar collagen content and organization without observer interpretation biases of staining intensity.
Extensive Applications Across Diseases
Fibrotic Diseases
- Liver Fibrosis
- Renal Fibrosis
- Pulmonary Fibrosis
- Bone Marrow Fibrosis
- Dermal Fibrosis
- Sub-epithelial Fibrosis on Cornea
- Retinal Pigment Epithelium
- Arterial Fibrosis
- Cardiac Fibrosis
- Biliary Disease
- Muscular Dystrophy (Sarcopenia)
- Alcoholic Liver Disease
- Alpha-1 antitrypsin Deficiency
- Biliary Atresia
Oncology
- Breast Cancer
- Pancreatic Cancer
- Liver Cancer
- Lung Cancer
- Ovarian Cancer
- Colon Cancer
- Kidney Cancer
- Bladder Cancer
- Squamous Cell Carcinoma of the Head and Neck (SCCHN)
Ushering in a New Era of Tissue Evaluation
Transforming Imaging in Pathology
Supplementing SHG microscopy with AI assessments, our platforms analyze liver biopsies to accurately quantify fibrosis – unlocking continuous metrics that sensitively detect subtle histological changes that may be missed by stains. The continuous values have shown good correlation with conventional pathology and objectively enhanced disease characterization, reduce variability in assessments, and provide greater sensitivity to accurately measure the magnitude of treatment efficacy.
Second Harmonic Generation FAQs
Second Harmonic Generation (SHG) imaging is a laser-based, nonlinear optical microscopy technique that captures high-contrast images of non-centrosymmetric structures in tissue, especially fibrillar collagen, without the need for chemical stains or dyes.
In SHG microscopy, tissue is illuminated with femtosecond laser pulses. When these pulses interact with structures like collagen fibers, two lower-energy photons combine to produce a new photon with twice the energy (half the wavelength). Special filters detect this signal to form a detailed image of tissue architecture.
Unlike traditional histology that relies on chemical stains like Masson’s trichrome or Picosirius red, SHG directly detects intrinsic optical signals from tissue components. This eliminates variability from staining protocols and produces consistent, reproducible imaging results with no photobleaching.
SHG is highly sensitive to fibrillar collagen, a key structural protein in many tissues. This makes it especially valuable for visualizing collagen organization in fibrotic diseases and evaluating structural changes in tissue extracellular matrix.
SHG imaging enables objective, quantitative analysis of fibrosis by visualizing collagen content and architecture in unstained tissue samples. This quantitative capability, particularly when combined with AI-based analysis, improves detection of subtle histological changes and supports more sensitive disease assessment than traditional staining.
- No chemical stains or dyes needed, reducing preparation variability.
- Non-destructive imaging, preserving samples for further analyses.
- Highly reproducible and quantitative, especially when combined with computational analysis.
Yes, when SHG image data is processed with AI and machine learning algorithms, it enables automated quantification of collagen morphology and fibrosis metrics, enhancing consistency, sensitivity, and clinical relevance in both research and therapeutic evaluation contexts.
While often used in liver fibrosis and MASH/MASLD research, SHG imaging also visualizes collagen in tissues like skin, lung, cornea, and other fibrotic or ECM-rich environments, making it broadly useful in biomedical research.