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. 2021 Jan 18;11:1693. doi: 10.1038/s41598-021-81465-4

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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FAP expression in CAFs and conjugation of FAP-IR700. (a) Immunofluorescence imaging of fibroblasts is shown. Normal FEF3 cells did not express FAP or αSMA, while CAFs stimulated with tumor cells expressed FAP on the cell surface and αSMA in the cytoplasm. Images are shown at 600 × magnification (green: FAP, red: αSMA, blue: nucleus). (b) WB demonstrated that the expression of αSMA was increased in CAFs compared to untreated FEF3 cells. (c) Flow cytometry demonstrated the expression of FAP. The relative mean fluorescence intensity of FAP was significantly increased in CAFs compared to untreated FEF3 cells as found by our FlowJo Software. Data are shown as the mean ± SD, Student’s t test. **P < 0.05. (d) The binding of FAP on CAFs and FAP-IR700 is shown by immunofluorescence imaging. After 6 h of conjugation with FAP-IR700, fibroblasts were fixed and observed by confocal microscopy. IR700 was observed in the same location as FAP expression in CAFs. However, it was not observed in normal FEF3 cells. Images are shown at 600 × magnification (green: FAP, blue: nucleus, magenta: IR700).