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High-Contrast Luminescent Immunohistochemistry Using PEGylated Lanthanide Complexes
Fei Su1,2, Xiongjian Luo1, Zhongbo Du1
1UTS-SUSTech Joint Research Centre for Biomedical Materials and Devices, Department of Biomedical Engineering, Southern University of Science and Technology, Shenzhen, Guangdong 518055, China.
Insights
This study enhances immunohistochemistry (IHC) contrast using bright, long-lifetime lanthanide probes and time-gated imaging. This method improves the detection of biomarkers like HER2 in tissue samples.
Area of Science:
- Biomedical imaging
- Molecular pathology
- Nanotechnology
Background:
- Immunohistochemistry (IHC) offers high resolution but suffers from limited contrast due to probe brightness and tissue autofluorescence.
- Autofluorescence in biological tissues significantly hinders the sensitivity of conventional fluorescent imaging techniques.
Purpose of the Study:
- To enhance imaging contrast in immunohistochemistry by developing novel high-density labeled lanthanide probes.
- To overcome the limitations of autofluorescence and improve the detection of low-abundance biomarkers in tissue sections.
Main Methods:
- High-density labeling of long-lifetime lanthanide complexes, specifically seven europium complexes conjugated to an eight-arm hydrophilic poly(ethylene glycol) (PEG) linker.
- Utilizing time-gated imaging to minimize background autofluorescence and amplify signal from the lanthanide probes.
- Demonstrating compatibility with tyramide signal amplification (TSA) for further signal enhancement.
Main Results:
- Achieved signal amplification and improved water solubility (up to 10 mg/mL) of the lanthanide probes due to PEGylation.
- Enhanced contrast by 2-fold for human epidermal growth factor receptor 2 (HER2) in FFPE tissues and 3-fold for cytokeratin 18 (CK18) in frozen sections.
- Successfully visualized biomarkers with improved sensitivity and reduced background noise.
Conclusions:
- The developed high-density labeled lanthanide probes combined with time-gated imaging significantly improve IHC contrast and sensitivity.
- This approach offers a promising strategy for the sensitive detection of low-abundance biomarkers in complex tissue environments.
- The method holds potential for advancing diagnostic capabilities in pathology and molecular imaging.
Abstract:
Immunohistochemistry (IHC) using fluorescent probes provides high resolution with multiplexing capability, but the imaging contrast is limited by the brightness of the fluorescent probe and the intrinsic autofluorescence background from tissues. Herein, we improved the contrast by high-density labeling of long-lifetime lanthanide complexes and time-gated imaging. As the large (∼280 nm) Stokes shift of lanthanide complexes effectively prevents the issue of concentration quenching, we succeeded in conjugating seven europium complexes to an eight-arm hydrophilic poly(ethylene glycol) (PEG) linker for signal amplification with improved water solubility to the level of up to 10 mg/mL. Moreover, we demonstrated that both human epidermal growth factor receptor 2 (HER2) in a formalin-fixed paraffin-embedded (FFPE) tissue section and cytokeratin 18 (CK18) in a frozen section can be resolved with the enhanced contrast by 2-fold and 3-fold, respectively. Furthermore, we show that the PEGylation of multiple lanthanide complexes is compatible with tyramide signal amplification (TSA). This work suggests new opportunities for sensitive imaging of low-abundance biomarkers in a tissue matrix.
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