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Cell-Penetrating Peptide Conjugations and Magnetic Cell Labels 339
Unlabled cells CLIO-Tat cells Mixed cells
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coated iron filings
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Fluorescence
FIGURE 15.7 Left: experimental set up. The FACS graphs from the three different cell
populations are shown on the right and bottom. Top right: mixed cells before separation;
bottom right: labeled cells after separation; bottom left: supernatant after separation. Separated
cells were 96% pure.
Our data show that CLIO–Tat-labeled lymphocytes were 96.8% pure after the
first separation cycle (approximately 1 min). The parent cell solution (supernatant)
was depleted from labeled lymphocytes from 43.4 to 17.5% during the first pass.
The mean fluorescence of CLIO–Tat-labeled cells was similar before and after
separation (221.8 ± 7.7 vs. 206.1 ± 8.6 AU respectively). Lymphocytes labeled with
non-labeled CLIO by fluid phase pinocytosis (20 ng Fe/million cells), could not be
extracted with the separator, presumably because the internalized amount was too
low. Virtually identical results were obtained when CD34+ cells were labeled with
CLIO–Tat; using a 50/50% mixture of CLIO–Tat to unlabeled CD34+, a single
magnetic separation yielded 98.95% pure CLIO–Tat-labeled CD34+ cells.
15.3.1.5 Recovery of In Vivo Injected Cells
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We subsequently hypothesized that CLIO–Tat-labeled cells could be recovered after
i.v. injection and in vivo homing. We therefore labeled mouse lymphocytes with
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Mixed cells before seperation
Retained cells
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