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Alexa fluor 700 anti human cd3

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Alexa Fluor® 700 anti-human CD3 is a fluorescently-labeled antibody that binds to the CD3 cell surface receptor on human T cells. This antibody can be used in flow cytometry applications to identify and quantify T cell populations.

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Lab products found in correlation

Fitc anti human tcrαβ, by BioLegend (2 mentions) Facsdiva, by FlowJo (1 mentions) Lsrfortessa, by BD (1 mentions) Facsdiva, by BD (1 mentions) Lsrfortessa cell analyzer, by BD (1 mentions) Apc anti human cd4, by BioLegend (1 mentions) Anti human cd69 pe, by BioLegend (1 mentions) Fitc anti human cd14, by BioLegend (1 mentions) Fitc anti human cd123, by BioLegend (1 mentions) Fitc anti human cd34, by BioLegend (1 mentions) Apc anti human cd336 nkp44, by BioLegend (1 mentions) Fitc anti human tcrγδ, by BioLegend (1 mentions) Alexa fluor 488 anti human cd19, by BioLegend (1 mentions) Fitc anti human fcεriα, by BioLegend (1 mentions) Pe anti human cd25, by BioLegend (1 mentions) Percp cyanine5.5 anti human cd8, by BioLegend (1 mentions) Facsaria 3, by BD (1 mentions) Facscelesta flow cytometer, by BD (1 mentions)

Close spelling variants of this product

Alexa Fluor 700-anti-CD3 CD3-Alexa Fluor 700 Alexa Fluor 700 Anti-human CD3 Anti-CD3

5 protocols using alexa fluor 700 anti human cd3

1

Purification of CD4+CD25- T Cells

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CD4 + CD25‐T (mentioned as CD4+ T cells in the manuscript for convenience unless otherwise specified) cells were purified from the PBMCs using a CD4 + CD25+ T cell isolation kit based on a two‐step negative isolation protocol, as per the manufacturer's prescription (Stem cell technologies). The kit separates CD4 + CD25+ and CD4 + CD25‐ T cells using positive and negative selection respectively. We collected negatively selected cell population and assessed cellular purity by immunophenotyping using anti‐human‐CD3 Alexa Fluor 700 (Clone OKT3), CD4 APC‐Cy7 (clone RPA‐T4), CD25 PE‐Cy7 (clone BC96) (Biolegend Inc., San Diego, CA, USA), and FoxP3‐BD Horizon V450 (Clone 236A/E7) (BD) antibodies on a flow cytometer, (LSR™ Fortessa, BD Biosciences, San Jose, CA, USA). Purity of isolated cells (CD3 + CD4 + CD25‐FoxP3‐) ranged between 90‐98% (Figure S1A‐C). All data were acquired on the same flow cytometer using FACS Diva™ software version 7.0 (BD Biosciences) and analysis was done by FACS Diva™ or Flowjo™ version 10.5.3 9.0 (Flowjo LLC, Ashland, OR, USA).
Sharma R.K., Sharma J., Kumar R., Badal D., Pattekar A., Sehgal S., Gupta A., Jain P, & Sachdeva N. (2022). TLR9 signalling activation via direct ligation and its functional consequences in CD4 + T cells. Scandinavian Journal of Immunology, 96(5), e13214.
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2

Isolation and Phenotyping of T Cells and Dendritic Cells

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The islets were removed from each co-culture by hand-picking. The cells were then administered into 5 mL polystyrene tubes through 70 μM filters. The T cells were stained with anti-human CD3 Alexa Fluor 700, anti-human CD4 APC, anti-human CD8 APC-Cy7, anti-human CD69 PE, and anti-human CD25 BV786 (Biolegend, San Diego, CA, USA). The dendritic cells were stained with anti-human CD14 Brilliant Violet 605, anti-human CD11c PE-Cy7, anti-human CD80 BUV737, and anti-human CD83 BV785 (Biolegend, CA, USA). The cells were then analyzed on a BD LSRFortessa Cell Analyzer.
Mattke J., Darden C.M., Vasu S., Lawrence M.C., Kirkland J., Kane R.R, & Naziruddin B. (2024). Inhibition of Toll-like Receptor 4 Using Small Molecule, TAK-242, Protects Islets from Innate Immune Responses. Cells, 13(5), 416.
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3

Multiparametric Flow Cytometry Panel

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Alexa Fluor® 700 anti-human CD3, (clone: HIT3a), FITC-anti human TCRαβ (clone: IP26), FITC-anti-human TCRγδ (clone: B1), APC/Cy7 Anti-Human CD127 (I-7Ra), (clone: A019D5), PE anti-human CD161, (clone: HP-3G10), Brilliant Violet 421TM anti-human CD117 (c-kit), (clone 104D2), PE/Cy7 anti-human CD294 (CRTH2), (clone: BM16), APC anti-human CD336 (NKp44), (clone: 325110), Alexa Fluor® 488 anti-human CD19, (clone: HIB19), FITC anti-human CD94, (clone:DX22), FITC anti-human CD1a, (clone: HI149), FITC anti-human CD11c, (clone: 3.9), FITC anti-human CD123, (clone: 6H6), anti-human CD303 (BDCA-2), (clone:201A), FITC anti-human CD14, (clone: 63D3), FITC anti-human FcεRIα, (clone: NP4D6), FITC anti-human CD34,(clone: 561), APC/Cy7 anti-human IFN-γ, (clone: 4S.B3) all from BioLegend. Anti-Human CD363 (S1PR1) eFluor® 660, (clone: SW4GYPP, ThermoFisher), Mouse IgG1 K Isotype Control eFluor® 660, (clone: P3.6.2.8.1, ThermoFisher). Anti-mouse CD3) APC, (clone: 17A2), anti-mouse NK1.1 Alexa Fluor® 647, (clone: PK136), anti-mouse B220 APC/Cy7 or FITC, (clone:RA3-6B2), anti-mouse CD45-Percp Cy5.5, (clone:30-F11), anti-mouse CD90.2 PE Cy7, (clone:30-H12), anti-mouse CD11b APC, (clone:M1/70), anti-mouse GATA3 PE (Clone: 16E10A23), anti-mouse Rorγt (Clone: 2F7-2).
Eken A., Yetkin M.F., Vural A., Okus F.Z., Erdem S., Azizoglu Z.B., Haliloglu Y., Cakir M., Turkoglu E.M., Kilic O., Kara I., Dönmez Altuntaş H., Oukka M., Kutuk M.S., Mirza M, & Canatan H. (2019). Fingolimod Alters Tissue Distribution and Cytokine Production of Human and Murine Innate Lymphoid Cells. Frontiers in Immunology, 10, 217.
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4

Single-Cell ATAC-Seq of Human PBMCs

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Human PBMCs were thawed after storage in liquid nitrogen and allowed to recover overnight in R10 culture medium (RPMI1640 medium supplemented with 10% fetal calf serum (FCS), 25 mM HEPES, 1* nonessential amino acids, 50 μM β-mercaptoethanol, and 1 × penicillin and streptomycin). Cell surface staining was performed with the following antibodies: DAPI, Alexa Fluor® 700 anti-human CD3 (Clone SK7, 1:100, BioLegend, Cat #344822), FITC anti-human TCR α/β (Clone IP26, 1:50, BioLegend, Cat #306706), PerCP/Cyanine5.5 anti-human CD8 (Clone SK1, 1:200, BioLegend, Cat #344710), PE/Dazzle™ 594 anti-human CD4 (Clone A161A1, 1:200, BioLegend, Cat #357411), PE anti-human CD25 (Clone BC96, 1:200, BioLegend, Cat #302606), Brilliant Violet 650™ anti-human CD127 (Clone A019D5, 1:200, BioLegend, Cat #351326), APC anti-human CD137 (4– 1BB) (Clone 4B4–1, 1:100, BioLegend, Cat #309810), Brilliant Violet 605 anti-human CD279 (PD-1) (Clone EH12.2H7, 1:100, BioLegend, Cat #329923). Single viable DAPI-CD3+TCRaß+ T cells were immediately indexed sorted into 96-well plates for Ti-ATAC-seq analysis using BD FACS Aria III.
You M., Chen L., Zhang D., Zhao P., Chen Z., Qin E.Q., Gao Y., Davis M.M, & Yang P. (2021). Single-cell epigenomic landscape of peripheral immune cells reveals establishment of trained immunity in individuals convalescing from COVID-19. Nature cell biology, 23(6), 620-630.
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5

Alveolus Lung-Chip PBMC Isolation and Characterization

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PBMC were harvested from the Alveolus Lung-Chip epithelial channel at T=48 hr (terminal timepoint) after dosing with PBMC-TCB, by repeated washing using 200 µL tips by blocking the chip inlet. PBMC from each chip was transferred to a V bottom 96-well plate and washed with DPBS + 1% v/v FBS solution before staining with surface markers. Master mix of surface markers was prepared in Brilliant Buffer solution (BD BioSciences) which consisted of anti-human Alexa Fluor 700 anti-human CD3 (BioLegend, cat. 300324), Brilliant Violet 785 anti-human CD4 (BioLegend, cat. 317442) and Brilliant Violet 650 anti-human CD69 (BioLegend, cat. 310934). Harvested PBMC was stained with the prepared master mix for 20 min at 4°C and fixed using 1% v/v paraformaldehyde in DPBS for 15 min at room temperature. Samples collected were then washed with DPBS + 1% v/v FBS solution and stored in 4°C and read within 3 days.
Sample data was acquired using BD FACSCelestaTM flow cytometer (BD BioSciences) and data was analyzed using FlowJo V10 software (FlowJo).
Kerns S.J., Belgur C., Petropolis D., Kanellias M., Barrile R., Sam J., Weinzierl T., Fauti T., Freimoser-Grundschober A., Eckmann J., Hage C., Geiger M., Ng P.R., Tien-Street W., Manatakis D.V., Micallef V., Gerard R., Bscheider M., Breous-Nystrom E., Schneider A., Giusti A.M., Bertinetti-Lapatki C., Grant H.S., Roth A.B., Hamilton G.A., Singer T., Karalis K., Moisan A., Bruenker P., Klein C., Bacac M., Gjorevski N, & Cabon L. (2021). Human immunocompetent Organ-on-Chip platforms allow safety profiling of tumor-targeted T-cell bispecific antibodies. eLife, 10, e67106.
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