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<h2>Characterization</h2>
 
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<h3>HiBiT Export</h3>
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We can successfully secrete our CD63-containing exosomes from HEK293T cells 48h after transfection as shown in Figure 3. Expression levels of exosomes containing the adapter L7Ae are around 150 pmol HiBiT-equivalent. Most importantly, the secretion efficiency of loaded exosomes is significantly higher than for control samples lacking the CD63-construct and even twice as high as the secretion efficiency of empty exosomes lacking the RNA binding protein (RBP).
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                <img src="" width="400" height="360" class="figure-img img-fluid rounded" alt="Secretion efficiency of exosome vesicles calculated from HiBiT-measurements.">
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Figure 3: Secretion efficiency of exosome vesicles calculated from HiBiT-measurements.</b> Loaded vesicles containing the adapter L7Ae as well as empty exosomes lacking the RNA binding protein (RBP) were secreted from HEK293T cells with efficiencies of more than 100 pmol HiBiT-equivalent. In contrast, control samples transfected with mock DNA did not show any secretion. Measurements were performed for n = 6 biological replicates in a 96-well format.
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Revision as of 15:11, 21 October 2019


CD63

CD63 is a tetraspanin found in exosomes. It is a cell surface receptor for Timp-1 and plays a role in the activation of cellular signaling cascades. CD63 can be used to load RNA or proteins into extracellular vesicles.


Usage and Biology

CD63 is a Tetraspanine. Tetraspanines are a superfamily of cell surface associated membrane proteins with four transmembrane domains. CD63 was the first characterized Tetraspanine. It is abundantly present in late endosomes and lysosomes as well as exosomes. The gene is located on the human chromosome 12q13. Although the intracellular function of CD63 remains to be established, a number of studies performed in different cell types implicate a role for CD63 in intracellular transport of other proteins[1]

Characterization

HiBiT Export

We can successfully secrete our CD63-containing exosomes from HEK293T cells 48h after transfection as shown in Figure 3. Expression levels of exosomes containing the adapter L7Ae are around 150 pmol HiBiT-equivalent. Most importantly, the secretion efficiency of loaded exosomes is significantly higher than for control samples lacking the CD63-construct and even twice as high as the secretion efficiency of empty exosomes lacking the RNA binding protein (RBP).

Secretion efficiency of exosome vesicles calculated from HiBiT-measurements.
Figure 3: Secretion efficiency of exosome vesicles calculated from HiBiT-measurements. Loaded vesicles containing the adapter L7Ae as well as empty exosomes lacking the RNA binding protein (RBP) were secreted from HEK293T cells with efficiencies of more than 100 pmol HiBiT-equivalent. In contrast, control samples transfected with mock DNA did not show any secretion. Measurements were performed for n = 6 biological replicates in a 96-well format.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]

References

  1. ↑ Trafficking and function of the tetraspanin CD63 Cell Microscopy Center, Department of Cell Biology and Institute of Biomembranes, University Medical Center Utrecht, Heidelberglaan 100, 3584CX Utrecht, The Netherlands Received 16 September 2008, Accepted 23 September 2008, Available online 7 October 2008.