2. Academic Validation
  3. Genetically encoded biosensor for monitoring spatiotemporal dynamics of CCR2 ligands in culture and in vivo

Genetically encoded biosensor for monitoring spatiotemporal dynamics of CCR2 ligands in culture and in vivo

  • Nat Methods. 2025 Aug;22(8):1731-1741. doi: 10.1038/s41592-025-02742-y.
Xian Xiao 1 2 3,  Chenyu Wang # 4 5 6,  Xun Guo # 1 2 3,  Fengxue Xi 4 5 6,  Qiuling Qian 1 2 3,  Guoteng Liang 1 2 3,  Ming Chen 7,  Xiaoting Sun 1 2 3,  Balint Szabo 8 9,  Miao Jing 5 6,  Kiryl D Piatkevich 10 11 12
Affiliations

Affiliations

  • 1 School of Life Sciences, Westlake University, Hangzhou, China.
  • 2 Westlake Laboratory of Life Sciences and Biomedicine, Hangzhou, China.
  • 3 Institute of Basic Medical Sciences, Westlake Institute for Advanced Study, Hangzhou, China.
  • 4 Basic Medical Sciences, Capital Medical University, Beijing, China.
  • 5 Beijing Institute for Brain Research, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, China.
  • 6 Chinese Institute for Brain Research, Beijing, China.
  • 7 Lingang Laboratory, Shanghai, China.
  • 8 CellSorter KFT, Budapest, Hungary.
  • 9 Eötvös University (ELTE), Department of Biological Physics, Budapest, Hungary.
  • 10 School of Life Sciences, Westlake University, Hangzhou, China. kiryl.piatkevich@westlake.edu.cn.
  • 11 Westlake Laboratory of Life Sciences and Biomedicine, Hangzhou, China. kiryl.piatkevich@westlake.edu.cn.
  • 12 Institute of Basic Medical Sciences, Westlake Institute for Advanced Study, Hangzhou, China. kiryl.piatkevich@westlake.edu.cn.
  • # Contributed equally.
PMID: 40665002 DOI: 10.1038/s41592-025-02742-y
Abstract

Chemokines regulate immune cell migration in development, homeostasis and inflammation, but the precise spatiotemporal pattern of chemokine release in vivo remains elusive due to the constraints of existing detection methodologies. Here, we report the engineering and characterization of a genetically encoded green fluorescent chemokine sensor, named CRAFi-CCR2, which utilizes the CCR2 receptor as a sensing moiety. In astrocytes, hCRAFi-CCR2, derived from the human CCR2B receptor, exhibited ~300% increase in fluorescence in response to mCCL2, with nanomolar affinity (2.5 nM). Activation of hCRAFi-CCR2 did not affect downstream signaling pathways, such as calcium mobilization and receptor internalization. Using this sensor, we performed 17-20 h of real-time imaging to observe endogenous mCCL2 release under inflammatory conditions, both in Cell Culture and in mice. In mouse brain, we observed spatial heterogeneity of CCL2 signal response on a scale of about 20-50 µm, highlighting the complexity of the immune system's spatiotemporal signaling.

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