After overnight fasting, 5 mL peripheral blood samples were collected in BD Vacutainer® CPT™ Tubes (USA) from the subjects. The tubes were gently mixed upside down 10 times and stored vertically. Within two hours, the tubes were centrifugated at 1500 g, 20 min, and 25 °C. The mononuclear precursor cells were isolated, washed, and resuspended in PBS (Gibco, USA). The isolated cells were incubated with two kinds of monoclonal antibodies: PE-conjugated anti-human CD34 (BD, USA) and allophycocyanin-conjugated anti-human VEGFR-2 (R&D Systems, USA). The cell sorting was performed on a FACSAria III (BD Biosciences, USA) at 488 nm to obtain double-positive cells, CD34 + VEGFR-2 + cells [23 (link)].
Pe conjugated anti human cd34
Manufactured by BD
Sourced in United States
PE-conjugated anti-human CD34 is a flow cytometry reagent that binds to the CD34 antigen, a cell surface glycoprotein expressed on hematopoietic stem and progenitor cells.
Automatically generated - may contain errors
2 protocols using pe conjugated anti human cd34
1
Isolation of CD34+ VEGFR-2+ Cells
2
Immunophenotyping of Transplanted Cells
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Mice were euthanized at 19 weeks post-transplantation and whole femurs and spleens were removed. Femurs were flushed with Dulbecco’s phosphate buffer saline to remove marrow. Single cell suspensions of bone marrow and spleen cells were created and processed as previously described,35 (link) except that commercially available red blood cell lysis buffer was used as described above. Cells were counted and 2 × 105 cells/staining reaction were distributed to the wells of a 96-well round bottom tissue culture plate. Cells were stained with antibodies as previously described,35 (link) except that Phycoerythrin (PE) conjugated anti-human CD3 (Clone UCHT1), PE conjugated anti-human CD11b (Clone ICRF-44), PE conjugated anti-human CD34 (Clone 581), and PE conjugated mouse IgG1, k isotype control (Clone MOPC-21) antibodies (BD Biosciences) were also used. Cells were washed and analyzed by flow cytometry. Statistical significance was determined by performing t-tests.
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