Anti-mouse CD117 (c-kit)-APC (clone 2B8), anti-mouse Ly-6 A/EA (Sca-1)-PE/Cy7 (clone D7), biotin-conjugated anti-mouse CD4 (clone GK1.5), anti-mouse CD8 (clone 53–6.7), anti-mouse CD45R/B220 (cloneRA3-6B2), anti-mouse Ly6G/Gr-1 (clone RB6-8C5), anti-mouse CD11b (clone M1/70), anti-mouse Ter-119 (clone Ter-119), and APC-Cy7-conjugated streptavidin were obtained from eBioscience (San Diego, CA, USA). Anti-mouse CD45.1-FITC (clone A20, Ly5.1), anti-mouse CD45.2-PE (clone104, Ly5.2), anti-mouse Ly6G/Gr-1-PE/Cy7 (cloneRB6-8C5), anti-mouse CD45R/B220-PerCP (cloneRA3-6B2), anti-mouse CD11b-PE/Cy7 (cloneM1/70), anti-mouse CD3-APC (clone145-2C11), and streptavidin-PerCP (405213) were obtained from Biolegend (San Diego, CA, USA). Anti-mouse γH2AX and anti-mouse cleaved CASPASE 3 antibodies were obtained from Cell Signaling Technology (Danvers, MA, USA). Anti-mouse HO-1, anti-mouse NQO1, anti-mouse cytochrome C, anti-mouse β-ACTIN, anti-mouse NRF2, anti-mouse BAX, anti-mouse BAK, and anti-mouse BCL-XL antibodies were obtained from Abcam (Cambridge, UK). FITC-conjugated goat anti-rabbit/mouse antibodies were obtained from ZSGB-BIO (Beijing, China). Cytofix/Cytoperm buffer and Perm/Wash buffer were obtained from BD Pharmingen (San Diego, CA, USA). ATX was obtained from Aladdin Co. (Shanghai, China).
Apc cy7 conjugated streptavidin
Manufactured by Thermo Fisher Scientific
Sourced in United States
APC-Cy7-conjugated streptavidin is a fluorescent label used in flow cytometry and other immunoassays. It consists of the streptavidin protein conjugated to the fluorescent dyes APC and Cy7. Streptavidin has a high affinity for biotin, allowing it to bind to biotinylated antibodies or other biomolecules. The APC-Cy7 fluorescent label can then be detected and quantified using appropriate instrumentation.
Automatically generated - may contain errors
Lab products found in correlation
Fitc conjugated goat anti rabbit antibody, by Abcam (3 mentions)
Cytofix cytoperm buffer, by BD (3 mentions)
Perm wash buffer, by BD (2 mentions)
Anti mouse ly 6a ea sca 1 pe cy7, by Thermo Fisher Scientific (2 mentions)
2 7 dichlorodihydrofluorescein diacetate dcf da, by Merck Group (2 mentions)
Ter119, by Thermo Fisher Scientific (2 mentions)
Cd45r b220, by Thermo Fisher Scientific (2 mentions)
M3434 methylcellulose, by STEMCELL (2 mentions)
Cd11b, by Thermo Fisher Scientific (2 mentions)
Fetal calf serum (fcs), by Sartorius (2 mentions)
Mouse anti β actin, by Abcam (1 mentions)
Anti mouse cleaved caspase 3, by Cell Signaling Technology (1 mentions)
Mouse anti γh2ax, by Cell Signaling Technology (1 mentions)
Mouse anti ho 1, by Abcam (1 mentions)
Mouse anti bax, by Abcam (1 mentions)
Rabbit anti γh2ax, by Cell Signaling Technology (1 mentions)
Anti mouse ly 6g ly 6c gr 1, by BD (1 mentions)
Anti mouse cd117 c kit apc, by Thermo Fisher Scientific (1 mentions)
Rabbit anti nox4, by Proteintech (1 mentions)
Cytoperm permeabilization buffer plus, by BD (1 mentions)
7 protocols using apc cy7 conjugated streptavidin
1
Multiparametric Immune Cell Analysis
2
Cellular Oxidative Stress Analysis
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XMU‐MP‐1 was provided by Selleck chemicals; 2′, 7′‐Di‐chlorodi‐hydrofluorescein diacetate (DCFDA) and dichloro‐fluorescein (DCF) were obtained from Sigma. Anti‐mouse Ly‐6G/Ly‐6C (Gr‐1) was abtained from BD bioscience. Anti‐mouse CD117(c‐kit)‐APC, anti‐mouse Ly‐6A/EA(Sca‐1)‐PE/Cy7, biotin‐conjugated anti‐mouse CD45R/B220, anti‐mouse CD11b, anti‐mouse Ter‐119 and APC‐Cy7‐conjugated streptavidin were obtained from eBioscience. Rabbit anti‐γ‐H2AX was obtained from Cell Signaling Technology. Rabbit anti‐NOX4 was obtained from Proteintech. FITC‐conjugated goat anti‐rabbit antibodies were obtained from Abcam Biotechnology. Cytofix/Cytoperm buffer, Perm/Wash buffer and Cytoperm Permeabilization Buffer Plus were obtained from BD Pharmingen.
3
Analyzing Hematopoietic Stem and Progenitor Cells
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BMMNCs were incubated with biotin-conjugated rat antibodies specific for murine CD5, Mac-1, CD45R/B220, Ter-119, and Gr-1 for 15 min at room temperature. After washing with PBS twice, the cells were stained with APC-Cy7-conjugated streptavidin,PE-Cy7-conjugated anti-Sca1, and Alexa Fluor 700-conjugated anti-c-kit antibodies (eBioscience, USA) and analyzed by flow cytometry. The ratio of HPCs (LSK-) and HSCs (LSK+) in BMMNCs was calculated.
4
Multicolor Flow Cytometric Analysis of Hematopoietic Cells
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Anti-mouse-CD45.1-FITC, CD45.2-PE, Gr-1-PE/CY7, CD45R/B220-PE/CY5.5 CD11b-PE/CY7 and CD3-APCwere purchased from BioLegend (San Diego, CA, USA); anti-mouse-Sca-1-PE, CD117 (c-kit) Alexa Fluor 700, CD4, CD8, CD45R/B220, Gr-1, CD11b, Ter119 and APC/CY7-conjugated streptavidin were purchased from eBioscience (San Diego, CA, USA); iron-dextrin was purchased from Pharmacosmos A/S (Denmark); the ROS staining kit (S0033) and NAC were purchased from the Beyotime Institute of Biotechnology; calcein-AM fluorescent dye was purchased from Sigma-Aldrich (USA); deferasirox was purchased from Novartis; RPMI 1640 was purchased from Gibco (USA); methylcellulose M3434 was purchased from Stem cell (USA); fetal calf serum was purchased from Bioind (Italy); CD117 MicroBeads (130-091-229) were purchased from Miltenyi Biotec (Germany); a RNA PCR Kit (AMV) Ver.3.0 (DRR019A) was purchased from Takara; a RNeasy MicroKit (74004) was purchased from Qiagen (Germany); the NOX4 and GPX1 gene qRT-PCR primers were synthesized by Sangon Biotech (Shanghai, China).
5
Multiparameter Immune Cell Analysis
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The anti-mouse Ly-6A/EA (Sca-1)-PE/Cy7, CD117 (c-kit), APC, biotin-conjugated CD5, CD4, CD8, CD45R/B220, Ly6G/Gr-1, CD11b, Ter-119, and APC/CY7-conjugated streptavidin antibodies were purchased from eBioscience (San Diego, CA, USA). The 21, 71-dichlorodihydrofluorescein diacetate (DCFDA) was purchased from Sigma-Aldrich (St Louis, MO, USA). The RPMI 1640 medium was purchased from Gibco (Grand Island, NY, USA). The BD Cytofix/Cytoperm buffer was purchased from BD Biosciences (San Diego, CA, USA). Methylcellulose M3434 was purchased from Stem Cell (Vancouer, BC, Canada). Fetal calf serum was purchased from Biological Industries (Kibbutz, Israel). The rabbit anti-H2AX antibody was obtained from Cell Signaling Technology (Danvers, MA, USA), the rabbit anti-NOX4 antibody from Proteintech (Wuhan, China) and the FITC-conjugated goat anti-rabbit antibodies from Abcam (Cambridge, MA, USA). Hei brick tea was purchased from Baishaxi Tea Industry (Hunan, China). Malondialdehyde (MDA), SOD, CATand GSHPx reagent kits were purchased from Nanjing Jiancheng Bioengineering (Jiangsu, China).
6
Murine Immune Cell Phenotyping and Oxidative Stress Analysis
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Eight- to 10-week-old male C57BL/6J mice were purchased from Vital River (Beijing, China), mice were housed in the Specific Pathogen Free level animal facility at the Institute of Radiation Medicine (IRM), the Chinese Academy of Medical Sciences (CAMS). All experimental procedures were approved by the Institutional Animal Care and Use Committee of the CAMS (Permit Number 1526, 7 April 2015), and written informed consent was obtained from all participants.
SB203580 was purchased from LC Laboratories (Boston, MA, USA). Biotin-conjugated anti-Mouse-CD4 (clone 34 GK1.5), anti-Mouse-CD8 (clone 53-6.7), anti-mouse-CD11b (clone M1/70), anti-mouse-CD45R/B220 (clone RA3-6B2), anti-mouse-Ly6G/Gr-1 (clone RB6-8C5), anti-mouse-Ter-119 (clone Ter-119), anti-mouse-CD117 (c-kit)-APC (clone 2B8), anti-mouse-Ly-6A/EA (Sca-1)-PE/Cy7 (clone D7), and APC-Cy7-conjugated streptavidin were purchased from eBioscience (San Diego, CA, USA). 2,7-dichlorodihydrofluorescein diacetate (DCFDA) was purchased from Sigma-Aldrich (St. Louis, MO, USA). Rabbit anti-p-p38 (ab61241), rabbit anti-p16, and FITC-conjugated goat anti-rabbit antibodies were obtained from Abcam Biotechnology (Cambridge, MA, USA).
SB203580 was purchased from LC Laboratories (Boston, MA, USA). Biotin-conjugated anti-Mouse-CD4 (clone 34 GK1.5), anti-Mouse-CD8 (clone 53-6.7), anti-mouse-CD11b (clone M1/70), anti-mouse-CD45R/B220 (clone RA3-6B2), anti-mouse-Ly6G/Gr-1 (clone RB6-8C5), anti-mouse-Ter-119 (clone Ter-119), anti-mouse-CD117 (c-kit)-APC (clone 2B8), anti-mouse-Ly-6A/EA (Sca-1)-PE/Cy7 (clone D7), and APC-Cy7-conjugated streptavidin were purchased from eBioscience (San Diego, CA, USA). 2,7-dichlorodihydrofluorescein diacetate (DCFDA) was purchased from Sigma-Aldrich (St. Louis, MO, USA). Rabbit anti-p-p38 (ab61241), rabbit anti-p16, and FITC-conjugated goat anti-rabbit antibodies were obtained from Abcam Biotechnology (Cambridge, MA, USA).
7
Isolation and Identification of Fetal Mouse LT-HSCs
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Fetal livers were isolated from mouse embryos at E14.5 under a dissecting microscope, and cell suspensions were prepared in PBS containing 2% FBS by repeatedly flushing through needles ranging from 18‐ to 27‐gauge. The cells were then passed through a nylon mesh with a pore size of 70 µm, and red blood cells were lysed using RBC lysis buffer (00‐4333‐57, eBioscience).
For staining LT‐HSCs, the cells were first incubated with Fc Block followed by biotin‐conjugated lineage marker antibodies (CD3e, CD4, CD5, CD8a, B220, Gr‐1, Ter119, and CD11b), followed by APC‐Cy7‐conjugated streptavidin, PE‐Cy7‐conjugated Sca‐1, APC‐conjugated CD11b, PE‐conjugated CD150, and Percp‐Cy5.5‐conjugated CD48 antibodies.[9 ] All antibodies were purchased from eBioscience or BioLegend. FACS data were analyzed using FlowJo software.
For staining LT‐HSCs, the cells were first incubated with Fc Block followed by biotin‐conjugated lineage marker antibodies (CD3e, CD4, CD5, CD8a, B220, Gr‐1, Ter119, and CD11b), followed by APC‐Cy7‐conjugated streptavidin, PE‐Cy7‐conjugated Sca‐1, APC‐conjugated CD11b, PE‐conjugated CD150, and Percp‐Cy5.5‐conjugated CD48 antibodies.[
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