Limits...
DNA modification of live cell surface.

Borisenko GG, Zaitseva MA, Chuvilin AN, Pozmogova GE - Nucleic Acids Res. (2009)

Bottom Line: By using fluorescence microscopy and flow cytometry we demonstrated that our synthetic conjugates of fatty acid with oligonucleotides can be incorporated in plasma membrane and then hybridized with complementary sequences at the cell surface.All procedures can be completed within minutes and do not alter cell viability.Using this approach we tethered floating myeloid HL-60 cells to adherent A431 epitheliocytes in a sequence specific fashion.

View Article: PubMed Central - PubMed

Affiliation: Research Institute of Physical-Chemical Medicine, Russian Academy of Science, Moscow 119312, Russia. grigoryb@yahoo.com

ABSTRACT
We report a novel approach for the attachment of DNA fragments to the surface of live cells. By using fluorescence microscopy and flow cytometry we demonstrated that our synthetic conjugates of fatty acid with oligonucleotides can be incorporated in plasma membrane and then hybridized with complementary sequences at the cell surface. Method permits to control amount of immobilized DNA on the cell surface. All procedures can be completed within minutes and do not alter cell viability. Using this approach we tethered floating myeloid HL-60 cells to adherent A431 epitheliocytes in a sequence specific fashion. Thus, this method allows rapid and simple DNA multicoding of the cell surface and, therefore, opens new opportunities in manipulating with cell-cell interactions.

Show MeSH

Related in: MedlinePlus

Fluorescence microscopy of Jurkat cells labeled with FT18NSte and FT18N. Cells were co-incubated with 0.2 μM FT18NSte (A, C) or FT18N (B, D) for 15 min at 37°C in PBS (green color). (C, D) Additionally, cells were labeled with 0.5 μM CTFR for 10 min at room temperature (red color). (E, F) Fluorescence profiles of single cells stained with CTFR and either FT18NSte (E) or FT18N (F); profiles were plotted via lines shown on panels C and D in violet color. (G, H) Cells were labeled with 5 µM PI for 5 min and with either FT18NSte (G) or FT18N (H). Dark spots on panel H represent unstained cells on the fluorescent background. Colocalization of green and red fluorescence is represented in the yellow color. (I) HL-60 cells were stained as in (C). (K) J774 cells were stained as in (A). (L) J774 cells were stained with 5 μM (Texas Red)-phosphatidylethanolamine. Photographs presented at panels K and L were taken at two focal plains: through the top (upper panel) and through the middle section of cells (bottom panel). The yellow marker size is 20 µm.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2651772&req=5

Figure 4: Fluorescence microscopy of Jurkat cells labeled with FT18NSte and FT18N. Cells were co-incubated with 0.2 μM FT18NSte (A, C) or FT18N (B, D) for 15 min at 37°C in PBS (green color). (C, D) Additionally, cells were labeled with 0.5 μM CTFR for 10 min at room temperature (red color). (E, F) Fluorescence profiles of single cells stained with CTFR and either FT18NSte (E) or FT18N (F); profiles were plotted via lines shown on panels C and D in violet color. (G, H) Cells were labeled with 5 µM PI for 5 min and with either FT18NSte (G) or FT18N (H). Dark spots on panel H represent unstained cells on the fluorescent background. Colocalization of green and red fluorescence is represented in the yellow color. (I) HL-60 cells were stained as in (C). (K) J774 cells were stained as in (A). (L) J774 cells were stained with 5 μM (Texas Red)-phosphatidylethanolamine. Photographs presented at panels K and L were taken at two focal plains: through the top (upper panel) and through the middle section of cells (bottom panel). The yellow marker size is 20 µm.

Mentions: Jurkat and HL-60 cells were co-incubated with 0.2 μM FT18NSte, a fatty acid derivative of FAM-labeled oligonucleotide. Fluorescence microscopic evaluation of cells revealed an efficient cell staining (Figure 4A and I). Co-staining of Jurkat cells with cell tracer CTFR, which covalently modifies intracellular proteins, further demonstrated that FT18NSte was localized in the PM (Figure 4C and E). In the presence of FAM-labeled oligonucleotide lacking fatty acid (FT18N) the majority of cells remained unstained, though few cells contained the stain, which was most likely localized in the nuclear region (Figure 4B, D and F).Figure 4.


DNA modification of live cell surface.

Borisenko GG, Zaitseva MA, Chuvilin AN, Pozmogova GE - Nucleic Acids Res. (2009)

Fluorescence microscopy of Jurkat cells labeled with FT18NSte and FT18N. Cells were co-incubated with 0.2 μM FT18NSte (A, C) or FT18N (B, D) for 15 min at 37°C in PBS (green color). (C, D) Additionally, cells were labeled with 0.5 μM CTFR for 10 min at room temperature (red color). (E, F) Fluorescence profiles of single cells stained with CTFR and either FT18NSte (E) or FT18N (F); profiles were plotted via lines shown on panels C and D in violet color. (G, H) Cells were labeled with 5 µM PI for 5 min and with either FT18NSte (G) or FT18N (H). Dark spots on panel H represent unstained cells on the fluorescent background. Colocalization of green and red fluorescence is represented in the yellow color. (I) HL-60 cells were stained as in (C). (K) J774 cells were stained as in (A). (L) J774 cells were stained with 5 μM (Texas Red)-phosphatidylethanolamine. Photographs presented at panels K and L were taken at two focal plains: through the top (upper panel) and through the middle section of cells (bottom panel). The yellow marker size is 20 µm.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2651772&req=5

Figure 4: Fluorescence microscopy of Jurkat cells labeled with FT18NSte and FT18N. Cells were co-incubated with 0.2 μM FT18NSte (A, C) or FT18N (B, D) for 15 min at 37°C in PBS (green color). (C, D) Additionally, cells were labeled with 0.5 μM CTFR for 10 min at room temperature (red color). (E, F) Fluorescence profiles of single cells stained with CTFR and either FT18NSte (E) or FT18N (F); profiles were plotted via lines shown on panels C and D in violet color. (G, H) Cells were labeled with 5 µM PI for 5 min and with either FT18NSte (G) or FT18N (H). Dark spots on panel H represent unstained cells on the fluorescent background. Colocalization of green and red fluorescence is represented in the yellow color. (I) HL-60 cells were stained as in (C). (K) J774 cells were stained as in (A). (L) J774 cells were stained with 5 μM (Texas Red)-phosphatidylethanolamine. Photographs presented at panels K and L were taken at two focal plains: through the top (upper panel) and through the middle section of cells (bottom panel). The yellow marker size is 20 µm.
Mentions: Jurkat and HL-60 cells were co-incubated with 0.2 μM FT18NSte, a fatty acid derivative of FAM-labeled oligonucleotide. Fluorescence microscopic evaluation of cells revealed an efficient cell staining (Figure 4A and I). Co-staining of Jurkat cells with cell tracer CTFR, which covalently modifies intracellular proteins, further demonstrated that FT18NSte was localized in the PM (Figure 4C and E). In the presence of FAM-labeled oligonucleotide lacking fatty acid (FT18N) the majority of cells remained unstained, though few cells contained the stain, which was most likely localized in the nuclear region (Figure 4B, D and F).Figure 4.

Bottom Line: By using fluorescence microscopy and flow cytometry we demonstrated that our synthetic conjugates of fatty acid with oligonucleotides can be incorporated in plasma membrane and then hybridized with complementary sequences at the cell surface.All procedures can be completed within minutes and do not alter cell viability.Using this approach we tethered floating myeloid HL-60 cells to adherent A431 epitheliocytes in a sequence specific fashion.

View Article: PubMed Central - PubMed

Affiliation: Research Institute of Physical-Chemical Medicine, Russian Academy of Science, Moscow 119312, Russia. grigoryb@yahoo.com

ABSTRACT
We report a novel approach for the attachment of DNA fragments to the surface of live cells. By using fluorescence microscopy and flow cytometry we demonstrated that our synthetic conjugates of fatty acid with oligonucleotides can be incorporated in plasma membrane and then hybridized with complementary sequences at the cell surface. Method permits to control amount of immobilized DNA on the cell surface. All procedures can be completed within minutes and do not alter cell viability. Using this approach we tethered floating myeloid HL-60 cells to adherent A431 epitheliocytes in a sequence specific fashion. Thus, this method allows rapid and simple DNA multicoding of the cell surface and, therefore, opens new opportunities in manipulating with cell-cell interactions.

Show MeSH
Related in: MedlinePlus