Cd38 bv421

Manufactured by BD

CD38-BV421 is a fluorescently labeled antibody that binds to the CD38 antigen. CD38 is a transmembrane glycoprotein expressed on the surface of various cell types, including lymphocytes, monocytes, and plasma cells. The BV421 fluorophore is used to label the CD38 antibody, enabling detection and analysis of CD38-positive cells through flow cytometry.

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4 protocols using cd38 bv421

Comprehensive Immune Profiling of PBMCs

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2 × 10⁶ PBMCs were stained with fixable blue dead cell stain (ThermoFisher) or Zombie NIR™ Fixable Viability Kit (Biolegend) in PBS, followed by washes and surface marker staining in Brilliant™ Stain Buffer (BD) with antibodies (Biolegend unless stated): CD3-BV786 (317330), CD4-BUV395 (BD) (563550), CD8-BV421 (301036), CD27-AF700 (356416), CD45RA-PE-Cy7 (304126), CD25-PEdazzle594 (356126), CD127-BV711 (351328), iTCR-PE (342904), CD19-AF488 (302219), IgD-BV510 (348220), HLA-DR-BV786 (307642), CD38-BV421 (356618), CD14-BV711 (301838), CD16-PEdazzle594 (302054), CD303-PerCP-Cy5.5 (354210) followed by subsequent washes and fixation in 2% PFA. Stains and washes were carried out in Biolegend cell staining buffer. Data was acquired using a BD LSRFORTESSA X-20 flow cytometer (1-2 × 10⁶ cells per sample) and analysis performed using FlowJo Single Cell Analysis Software (TreeStar). Cytometer Setup and Tracking (CS&T) (BD) beads were used to monitor cytometer performance. Application settings were applied prior to compensation to ensure that all immunophenotyping data was comparable over time. Gating strategies are shown in Supplementary Figure 1.

Robinson G.A., Waddington K.E., Coelewij L., Peng J., Naja M., Wincup C., Radziszewska A., Peckham H., Isenberg D.A., Ioannou Y., Ciurtin C., Pineda-Torra I, & Jury E.C. (2021). Increased apolipoprotein-B:A1 ratio predicts cardiometabolic risk in patients with juvenile onset SLE. EBioMedicine, 65, 103243.

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Multiparametric Flow Cytometric Immunophenotyping

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Peripheral blood mononuclear cells were washed in PBS with 1% BSA (bovine serum albumin) and incubated for 30 min at 25°C with fluorescently labeled antibodies specific for B and T cells. Subsequently, samples were centrifuged and resuspended in propidium iodide (PI) solution (1 μg/ml PI and 10 μg/ml RNase A in PBS) and analyzed using BD FACS Canto II flow cytometer (BD Biosciences). The B cell panel included the following antibodies: IgA FITC, IgD PE, CD3 PerCP-Cy5.5, CD27 APC-H7, CD19 PE-Cy7, β7 APC, and CD38 BV421 from BD Biosciences. The T cell panel included the following antibodies: CXCR5 BV421, CXCR3 PE, CD4 APC-H7, CD3 FITC, CD196 APC, CD279 (PD-1) BV510, and CD45RA PE-Cy7 from BD Biosciences. Results were analyzed using FACSDiva software (BD Biosciences) and reported as MFI, reflecting the levels of cell surface antigens and relative cell count with respect to hierarchically higher cell populations (%). Cellular aggregates were eliminated using morphology parameters (FSC-A and FSC-H) (see Supplementary Figures 1, 2 for gating strategy).

Lodde V., Floris M., Beerman I., Munk R., Guha R., Steri M., Orrù V., Abdelmohsen K., Crompton P.D., Gorospe M., Idda M.L, & Cucca F. (2020). Evolutionarily Selected Overexpression of the Cytokine BAFF Enhances Mucosal Immune Response Against P. falciparum. Frontiers in Immunology, 11, 575103.

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Plasmablast Formation from B Cells

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B cell stimulation with a minor cocktail of stimuli was performed to study the effect of IL-21, co-stimulation, and BCR activation on plasmablast formation. CD19⁺ B cells were isolated via CD43 negative selection with CD43 MicroBeads (Miltenyi Biotec, Bergisch Gladbach, Germany) (purities ≥85%). B cells were incubated with anti-IL-21R antibody ATR-107 (10 μg/ml, Pfizer) or isotype-matched control (10 μg/ml IgG1-Fc, R&D systems). Next, cells were stimulated with 5 μg/ml soluble anti-CD40 (Bioceros, Utrecht, The Netherlands), 10 μg/ml goat-anti-human IgM (Jackson Immunoresearch, West Grove, PA, USA) and human recombinant IL-21 (100 ng/ml, eBioscience). Subsequently, the presence of plasmablasts on day 0 and the differentiation of memory B cells into plasmablasts on day 8 were determined with flow cytometry. Plasmablasts were defined as CD19^posCD27^highCD38^high cells (16 (link)). The following MoAbs were used: CD19 BV510 (Biolegend), CD27 Pe-Cy7 (eBioscience), IgD APC-Cy7 (Biolegend), and CD38 BV421 (BD Biosciences). In addition, viability staining with 7-AAD PerCP was performed (BD Biosciences).

de Leur K., Dor F.J., Dieterich M., van der Laan L.J., Hendriks R.W, & Baan C.C. (2017). IL-21 Receptor Antagonist Inhibits Differentiation of B Cells toward Plasmablasts upon Alloantigen Stimulation. Frontiers in Immunology, 8, 306.

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Comprehensive Immunophenotyping Antibody Panel

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The following antibodies were used to define antigen expression by flow-cytometry:
CD82-PE (Biolegend, 342103); CD120b (TNF-RII)-PE (Miltenyi, 130–107-740); EMR2-FITC, -APC (Miltenyi, 130–104-654, 130–104-656); ITGB5-PE (Biolegend, 345203); CCR1-PE, -APC (Miltenyi, 130–100-367, 130–100-358); CD96-APC (Miltenyi, 130–101-031); PTPRJ (CD148)-PE (life technologies, A15799); CD70-PE, -FITC (Biolegend, 355104, 355106); CD85d (ILT4)-PE -APC (Miltenyi, 130–100-567, 130–100-559); LTB4R1-AF700 (Novus Biologicals, FAB099N); CD85h (ILT1)-APC (Miltenyi, 130–100-920); TLR2-APC (Miltenyi, 130–099-020); CR1 (aka CD35)-APC (Miltenyi, 130–099-923); ITGAX (CD11c)-APC (Biolegend, 301613); EMB (abcam, 179801); EMC10 (abcam PA5–25112); LILRB3-PE (Miltenyi, 130–101-662); LILRB4-APC (R&D, FAB24251A); DAGLB (abcam, PA5–26331); P2RY13 (Novus Biologicals, NBP2–37382); LILRA6-APC (Miltenyi, 130–101-665); SLC30A1 (Alomone labs, AZT-011); SLC6A6 (LSBio, LS-C179237); SEMA4A (R&D, FAB4694A); CD123-PE (BD Biosciences, 555644); CLEC12A-PE (Miltenyi, 130–106-482); CD33-APC (Miltenyi, 130–098-864); CD38-BV421 (BD Biosciences, 562444); CD34-PE/Cy7 (Biolegend, 343515); CD45RA-BV640 (Biolegend, 304135); CD90-FITC (BD Biosciences, 555595).

Perna F., Berman S., Soni R., Mansilla-Soto J., Eyquem J., Hamieh M., Hendrickson R., Brennan C, & Sadelain M. (2017). Integrating proteomics and transcriptomics for systematic combinatorial chimeric antigen receptor therapy of AML. Cancer cell, 32(4), 506-519.e5.

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