The hAMSCs were extracted as previous described [15 (link)]. Third-passage (P3) cells were harvested from phenotype identification through staining with antibodies against CD34 (581, BD Pharmingen, USA), CD45 (HI30, BD Pharmingen, USA), HLA-DR (G46-6, BD Pharmingen, USA), CD11b (ICRF44, BD Pharmingen, USA), CD90 (5E10, BD Pharmingen, USA), CD73 (AD2, BD Pharmingen, USA) and CD105 (SN6, ebioscience, USA), and then analyzed by flow cytometer (BD FACS Canto II). The cells were checked for their multilineage differentiation using a specific induction medium (BGscience, China), and stained with Oil Red-O and Alizarin Red for adipogenesis and osteogenic differentiation, respectively. hAMSCs at passages 3 to 6 were used for the experiments.
Cd34 581
Manufactured by BD
Sourced in United States
The CD34 (581) is a laboratory equipment product. It is used to detect and quantify the presence of CD34 cells in biological samples. CD34 is a cell surface glycoprotein that serves as a marker for hematopoietic stem and progenitor cells. The core function of the CD34 (581) is to facilitate the identification and analysis of CD34-positive cells.
Automatically generated - may contain errors
Lab products found in correlation
Cd90 5e10, by BD (3 mentions)
Mouse fc block, by BD (2 mentions)
Cd45 hi30, by Thermo Fisher Scientific (2 mentions)
Facsaria 2, by BD (2 mentions)
Diva software, by BD (2 mentions)
Sfemii media, by STEMCELL (2 mentions)
Cd133 2 293c3, by Miltenyi Biotec (2 mentions)
Facscanto 2, by BD (1 mentions)
Cd73 ad2, by BD (1 mentions)
Cd45 hi30, by BD (1 mentions)
Facscalibur, by BD (1 mentions)
Annexin 5, by BD (1 mentions)
6 protocols using cd34 581
1
Characterization of Human Amniotic Mesenchymal Stem Cells
2
Multilineage hematopoietic engraftment analysis in NSG mice
3
Flow Cytometric Immunophenotyping of Cells
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Flow cytometry was performed on FACSCalibur (BD Biosciences, Franklin Lakes, NJ, United States) and analyzed using FlowJo V10 software (Tree Star, Ashland, OR, United States). Relative fluorescence intensities were calculated from the MFI value of the relevant staining divided by the MFI value of the medium control sample. Fluorochrome-labeled monoclonal antibodies were anti-human CD11b (ICRF44), CD15 (HI98), CD33 (WM53), CD34 (581), CD80 (L307.4), CD83 (HB15e), CD86 (2331), CD117 (YB5.B8), CD274 (MIH1), HLA-class I (G46-2.6), HLA-DR (G46-6), and Annexin V (all BD Biosciences).
4
Isolation and Characterization of HSPC Subsets
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Mobilized blood or fetal liver–derived CD34+ HSPCs were treated with AAV6 vectors and ZFN mRNA as described above. One or two days post-mRNA electroporation, cells were washed, blocked in FCS (Denville), and stained with the following fluorophore conjugated antibodies: CD34 (581) (BD Biosciences), CD90 (5E10) (BD Biosciences), and CD133/2 (293C3) (Miltenyi Biotec). Cell sorting into subsets based on expression of these markers was performed using a BD FACS Aria II (BD Biosciences), with all compensations performed using Diva software (BD Biosciences). Subsets were defined as primitive (P; CD34+CD133+CD90+), early (E; CD34+CD133+CD90−), and committed (C; CD34+CD133−CD90−) progenitors23 (link). The subsets derived from fetal liver HSPCs were cultured in fetal liver maintenance media, and the subsets derived from mobilized blood HSPCs were maintained in SFEM-II media (Stemcell Technologies, Vancouver, Canada) supplemented with SCF, Flt3L, TPO, and IL-6, for a further 6–7 days. GFP expression was determined in each population by flow cytometry, and site-specific insertion of GFP at either the CCR5 or AAVS1 loci was analyzed by In-Out PCR, as described above.
5
Multilineage hematopoietic engraftment analysis in NSG mice
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NSG mice were sublethally irradiated (315 cGy) one day prior to intrafemoral injection with transduced cells carried in IMDM 1% FBS at 25μl per mouse. Injected mice were analyzed for human hematopoietic engraftment at 12-14 weeks post transplantation or 3 and 6.5 weeks for STRC experiments. Mouse bones (femurs, tibiae and pelvis) and spleen were harvested and bones were crushed with a mortar and pestle then filtered in to single cell suspensions. Bone marrow and spleen cells were blocked with mouse Fc block (BD Biosciences) and human IgG (Sigma) and then stained with fluorochrome-conjugated antibodies specific to human hematopoietic cells. For multilineage engraftment analysis, cells from mice were stained with CD45 (HI30) (Invitrogen), CD33 (P67.6), CD15 (HI98), CD14 (MφP9), CD19 (HIB19), CD235a/GlyA (GA-R2), CD41a (HIP8) and CD34 (581) (BD Biosciences).
6
Isolation and Characterization of HSPC Subsets
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