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Correlative light-electron microscopy (CLEM) combining live-cell imaging and immunolabeling of ultrathin cryosections

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Figure 1: Schematic overview of the CLEM procedure.
Figure 2: CLEM in live HepG2 cells.

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Acknowledgements

We thank M. v. Peski and R. Scriwanek for assistance with figure and movie preparations and all other colleagues of the Department of Cell Biology for fruitful discussions. We are grateful to R. Pepperkok (EMBL, Heidelberg) for introducing CR into live-cell imaging. We thank E. Dell'Angelica for the construct encoding LAMP-1–mGFP (University of California, Los Angeles). J.K. is the recipient of VICI grant 918.56.611 of the Netherlands Organization for Scientific research

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Authors and Affiliations

  1. Department of Cell Biology and Institute of Biomembranes, Cell Microscopy Center, University Medical Center Utrecht, Heidelberglaan 100, Utrecht, 3584, CX, The Netherlands

    Carolien van Rijnsoever, Viola Oorschot & Judith Klumperman

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  1. Carolien van Rijnsoever
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  2. Viola Oorschot
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  3. Judith Klumperman
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Correspondence to Judith Klumperman.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–2, Supplementary Table 1 (PDF 230 kb)

Supplementary Video 1

Time lapse movie showing endosomes moving with distinct dynamics. HepG2 cells were transfected with a LAMP-1-mGFP expressing construct10 and incubated with dextran and Tf-Alexa568 according to the protocol described in figure 2. LAMP-1-mGFP is present in the Golgi region and in endosomes. Note a ‘ring’ endosome that remains stationary in the Golgi region (first arrow), whereas an endosome loaded only with dextran and/or Tf-Alexa568 moves through the cytoplasm (second arrow). Still frames and CLEM images of the imaged cell are shown in figure 2. (MOV 5914 kb)

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van Rijnsoever, C., Oorschot, V. & Klumperman, J. Correlative light-electron microscopy (CLEM) combining live-cell imaging and immunolabeling of ultrathin cryosections. Nat Methods 5, 973–980 (2008). https://doi.org/10.1038/nmeth.1263

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