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Cd38 bv421

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CD38-BV421 is a fluorescently-labeled antibody that targets the CD38 cell surface molecule. It is a tool for the identification and analysis of CD38-expressing cells using flow cytometry.

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Lab products found in correlation

Cd3 fitc, by BioLegend (3 mentions) Lsrfortessa flow cytometer, by BD (2 mentions) Cd45ra af700, by BD (2 mentions) Tigit pe cy7, by Thermo Fisher Scientific (2 mentions) Cd160 af488, by BD (2 mentions) Tim 3 bv650, by BD (2 mentions) Cd28 bv711, by BioLegend (2 mentions) Cd27 bv650, by BioLegend (2 mentions) Lag 3 apc, by Thermo Fisher Scientific (2 mentions) Hla dr af700, by BioLegend (2 mentions) Ccr7 bv421, by BioLegend (2 mentions) Pd 1 bv711, by BD (2 mentions) Facscelesta, by BD (2 mentions) Cd8 apc, by BD (2 mentions) Cd4 pe cy7, by BioLegend (2 mentions) Hla dr pe, by BioLegend (2 mentions) Propidium iodide, by Merck Group (2 mentions) Moflo legacy sorter, by Beckman Coulter (2 mentions) Quick rna microprep lysis buffer, by Zymo Research (2 mentions) Cd45ra apc h7, by BD (1 mentions)

Spelling variants of this product

Anti-CD38-BV421 (4 citations) CD4 PE-Cy7 (SK3); CD8 PerCP-Cy5.5 (RPA-T8); CCR7 FITC (G043H7); CD45RA BV421 (HI100); HLA-DR PE (L243); CD38 BV605 (HIT2) (2 citations)

10 protocols using cd38 bv421

1

Multiparameter Flow Cytometry Phenotyping

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Standard extracellular staining (20 mins at 24°C) was performed using the following fluorophore-labeled antibodies: CD3-BUV737 (BD Biosciences), CD4-BUV395 (BD Biosciences), CD8-BV711 (BD Biosciences), CD14-BV510 (BD Biosciences), CD19-BV510 (BioLegend), Live/Dead Fixable Aqua (ThermoFisher), CCR7-PE-CF594 (BD Biosciences), CD45RA-APC-H7 (BD Biosciences), CD28-APC-R700 (BD Biosciences), CD57-PE (BioLegend), PD-1-BV605, BV421 (BioLegend), CD69-BV650 (BioLegend), CD38-BV421 (BioLegend), HLA-DR-PerCP-Cy5.5 (BD Biosciences), TIGIT-PE-Cy7 (BioLegend), DNAM-BB515 (BD Biosciences), FcγRIIB-BV786 (BD Biosciences). Fluorescence minus one (FMO) controls were used to determine negative expression of FcγRIIB. Apoptosis was measured with Caspase-3/7 Green Flow Cytometry Assay Kit (ThermoFisher) or FITC Annexin V with 7-AAD Viability Staining Solution (BioLegend), according to the manufacturer’s instructions.
Sun H., Hartigan C.R., Chen C.W., Sun Y., Tariq M., Robertson J.M., Krummey S.M., Mehta A.K, & Ford M.L. (2021). TIGIT regulates apoptosis of risky memory T cell subsets implicated in belatacept-resistant rejection. American journal of transplantation : official journal of the American Society of Transplantation and the American Society of Transplant Surgeons, 21(10), 3256-3267.
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2

Comprehensive Cytokine and T-cell Profiling

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Antibodies for intracellular cytokine staining included: anti-CD3-AF700, CD4-PcPCy5.5, CD8-PECF594, IFNg-FITC, CD14-V500, CD19-V500, TNFα-APC, CD38-PECy7, HLADR-BV605 (BD Biosciences); PD-1-PE, IL-2-BV421 (BioLegend); Live-Dead-AquaViD (LifeTechnologies).
The T regulatory cell/activation panel included: anti-CD3-AF700, CD25-PE, HLA-DR-FITC (BD); CD4-PETxR, CD8-APC AF750, Live-Dead-AquaViD (LifeTechnologies); CD39-PECy7, FOXP3-APC (eBioscience); CD127- PE Cy5.5 (Beckman Coulter); CD38- BV421 (BioLegend).
Barreto-Duarte B., Sterling T.R., Fiske C.T., Almeida A., Nochowicz C.H., Smith R.M., Barnett L., Warren C., Blackman A., Lapa e Silva J.R., Andrade B.B, & Kalams S.A. (2020). Increased Frequency of Memory CD4+ T-Cell Responses in Individuals With Previously Treated Extrapulmonary Tuberculosis. Frontiers in Immunology, 11, 605338.
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3

Multiparametric Flow Cytometry of PBMCs

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PBMCs were incubated with directly conjugated antibodies for 30 min at 4°C. The cells were washed before flow cytometry analysis. Antibodies used included anti-human CD3-BV786, CD4-APC-Fire750, CD8-BV510, CD45RA-AF700, CD70-PE, PD-1-BV711, 2B4-FITC, CD160-AF488, TIM-3-BV650, CD95-PE-CY7 (BD Biosciences, San Diego, CA, USA), CCR7-BV421, HLA-DR-AF700, CD38-BV421, CD28-BV711, CD27-BV650 (BioLegend, San Diego, CA, USA), TIGIT-PE-Cy7, LAG-3-APC (Ebioscience, San Diego, CA, USA) and the corresponding isotype controls. Data acquisition was performed on an LSR Fortessa flow cytometer (BD Biosciences), and data was analyzed with FlowJo software (Tree Star, Ashland, OR, USA).
Wang D., Du J., Song Y., Wang B., Song R., Hao Y., Zeng Y., Xiao J., Zheng H., Zeng H., Zhao H, & Kong Y. (2020). CD70 contributes to age-associated T cell defects and overwhelming inflammatory responses. Aging (Albany NY), 12(12), 12032-12050.
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4

Comprehensive Immunophenotyping Panel

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Anti‐human GM‐CSF‐PE, CD3‐APC, CD33‐PE (Miltenyi Biotec, Auburn, CA); CD45RA‐Pacific blue, CCR7‐APC/cy7, CD4‐Pacific blue, CD8‐APC/cy7, CD19‐APC‐Cy7, CD3‐FITC, CD11b‐APC, CD38‐BV421, CD33‐BV605, CD34‐APC, CD45RA‐BV786 and CD16‐Pacific Blue (Biolegend, San Diego, CA); and CD116‐PE (BD Biosciences, Franklin Lakes, NJ) antibodies were used for analysis. Flow cytometric data were acquired by BD FACSCanto™II, BD FACSCelesta™ or BD Accuri™ C6 Plus (BD Biosciences San Jose, CA) and analysed by FlowJo (TOMY Digital Biology, Tokyo, Japan).
Hasegawa A., Saito S., Narimatsu S., Nakano S., Nagai M., Ohnota H., Inada Y., Morokawa H., Nakashima I., Morita D., Ide Y., Matsuda K., Tashiro H., Yagyu S., Tanaka M, & Nakazawa Y. (2021). Mutated GM‐CSF‐based CAR‐T cells targeting CD116/CD131 complexes exhibit enhanced anti‐tumor effects against acute myeloid leukaemia. Clinical & Translational Immunology, 10(5), e1282.
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5

Circulating Tumor Plasma Cell Detection

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Circulating tumor plasma cells in the peripheral blood (PB) were detected using flow cytometry. Briefly, PB samples were collected in tubes containing EDTA and processed using a bulk-lysis procedure (Versalyse, Beckman Coulter, Brea, CA, USA) within 24 h of sample collection. Labeling was performed using a single 8-colors antibody combination: CD45-KrOr, CD138-PC5.5, CD56-AA700, CD27-PC7, CD19-ECD, CD200-AA750 (Beckman Coulter), Kappa-APC, and Lambda FITC (BD Biosciences, Franklin Lakes, NJ, USA), and CD38-BV421 (Biolegend, San Diego, CA, USA). First, membrane staining was performed and this was followed after a PBS washing procedure by intracytoplasmic labeling (anti-Kappa and anti-Lambda) using a Fix & Perm Cell Permeabilization Kit (Beckman Coulter, Brea, CA, USA). After the final PBS wash, the final cell pellet was resuspended in 500 µL of PBS before analysis using the flow cytometer (Navios, Beckman Coulter, Brea, CA, USA). From 200,000 to 1 million leucocytes were acquired for each patient to ensure the quality of the analysis and to allow a limit of detection to be set at ≥20 tumor plasma cells (0.01% to 0.002%, according to the number of leucocytes). Data were analyzed using Kaluza 2.1 software (Beckman Coulter, Brea, CA, USA).
Fouquet G., Wartski M., Dechmi A., Willems L., Deau-Fischer B., Franchi P., Descroocq J., Deschamps P., Blanc-Autran E., Clerc J., Bouscary D., Barreau S., Chapuis N., Vignon M, & Cottereau A.S. (2021). Prognostic Value of FDG-PET/CT Parameters in Patients with Relapse/Refractory Multiple Myeloma before Anti-CD38 Based Therapy. Cancers, 13(17), 4323.
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6

Multiparametric Immune Profiling of PBMC

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Cryopreserved PBMCs were thawed and left for 1 h at 37 °C in the CO2 incubator. Subsequently the cells were collected and stained with BD Horizon™ Fixable Viability Stain 510 to identify live cells, as per manufacture protocol. The cells were then washed, and surface stained for IL-21R PE expression on, (i) CD4, CD8 and Tfh (CD4/CD45RA -/CXCR5+/PD1+); (ii) B cell CD20, B1(CD20+CD27+CD43+), Plasma blast (CD20+CD38+) and iii) monocytes CD14+HLADR+.
After staining, the cells were washed and fixed using 2% PFA. Acquisition was done on BD FACSCelesta (Becton-Dickenson, San Jose, CA). Forward and side scatters and singlets were used to gate and exclude cellular debris. The flow cytometry results were analyzed using FlowJo™ v10.8 Software (BD Life Sciences, Ashland, OR). The details of the antibodies used are as follows: Fixable Viability stain510, CD4 FITC (clone: RPA-T4) from BD Bioscience (San Jose, CA), IL-21R PE (clone: 2G1-K12), CD8 PerCP (clone: SK1), PD1 APC (clone: EH12.2H7), CXCR5 BV421 (clone: J252D4), CD45RA BV605 (clone: HI100), CD38 BV421 (clone: HB-7), CD16 AF700 (clone: B73.1), CD14 BV650 (clone: M5E2), HLADR BV605 (clone: L243), CD20 PerCP (clone: 2H7), CD27 FITC (clone: M-T271), CD43 APC (clone: CD43-10G7) from BioLegend (San Diego, CA).
Agrawal S., Baulch J.E., Madan S., Salah S., Cheeks S.N., Krattli RP J.r., Subramanian V.S., Acharya M.M, & Agrawal A. (2022). Impact of IL-21-associated peripheral and brain crosstalk on the Alzheimer’s disease neuropathology. Cellular and Molecular Life Sciences, 79(6), 331.
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7

Flow Cytometry Sorting of PBMCs

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PBMCs were separated from whole blood and frozen. Freshly thawed cells were washed and incubated with CD3-FITC (Biolegend), CD4-PECy7 (Biolegend), CD8-APC (BD Biosciences), HLA-DR-PE (Biolegend), and CD38-BV421 (Biolegend) for 40 minutes at 4°C per the manufacturer’s recommendation. Immediately before sorting, plasma membrane compromised cells were labeled with propidium iodide (Sigma). Fluorescence-activated cell sorting (FACS) was performed on a MoFlo Legacy Sorter (Beckman-Coulter) at the Johns Hopkins School of Public Health Flow Cytometry Core Facility. Gates were set using fluorescence minus one plus isotype control. Because flow and sorting had to be performed over several sessions, the gates were set each session using a reference participant’s PBMCs (selected based on availability of cells). The population of interest was sorted directly into ≥4 volumes of Quick-RNA® MicroPrep lysis buffer (Zymo Research) per the manufacturer’s recommendation. Sorting was stopped when the number of sorted cells reached 125,000 cells, although many samples did not reach this number. Flow cytometry analysis on two randomly selected post-sort samples revealed >95% purity. Sorted samples were vortexed, incubated for 10 minutes at room temperature, vortexed again, and frozen at −80°C until isolation.
EL-DIWANY R., BREITWIESER F.P., SOLIMAN M., SKAIST A.M., SRIKRISHNA G., BLANKSON J.N., RAY S.C., WHEELAN S.J., THOMAS D.L, & BALAGOPAL A. (2017). Intracellular HIV-1 RNA and CD4+ T cell Activation in Patients Starting Antiretrovirals. AIDS (London, England), 31(10), 1405-1414.
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8

Multiparametric Flow Cytometry Analysis

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Following collection of patient specimens, the following subsets were analyzed by FCM: whole BM, BMNCs, CD138+ cells, and CD138− cells. For each sample, at least 200,000 cells were washed and incubated for 30 min at 4°C with CD138-PeCy7, CD38-BV421, CD45-APC, CD19-PE, and CD56-FITC, or matching isotype controls (BioLegend), at a 1:200 dilution. Stained cells were washed once and fixed with 1% paraformaldehyde (PFA). For analysis of previously frozen primary CD138+ and CD138− cells, BMNCs, T cells, and B cells, Human TruStain FcX Fc Receptor Blocking Solution (BioLegend) was used according to the manufacturer’s directions. Cells were then stained with CD5-BV421, CD147-PE/Cy7, CD166-PE, CD205-APC (BioLegend), and CD98hc-FITC (ThermoFisher), or matching isotype controls (BioLegend). All samples were analyzed on a BD LSRFortessa X-20 flow cytometer (BD Biosciences). Data were analyzed using FlowJo software (FlowJo LLC).
Oldham R.A., Faber M.L., Keppel T.R., Buchberger A.R., Waas M., Hari P., Gundry R.L, & Medin J.A. (2020). Discovery and validation of surface N-glycoproteins in MM cell lines and patient samples uncovers immunotherapy targets. Journal for Immunotherapy of Cancer, 8(2), e000915.
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9

Comprehensive Immune Phenotyping of PBMCs

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PBMCs from healthy subjects were resuspended in PBS buffer and were incubated with directly conjugated antibodies for 30 min at 4°C. The cells were washed with 1× PBS before flow cytometry analysis. Antibodies used included anti-human CD3-BV786 or CD3-BUV737, CD8-BV510 or CD8-BUV395, CD160-AF488, CD45RA-AF700, CD28-APC, CD95-PE, CD57-BV421, PD-1-BV711, TIM-3-BV650 (BD Biosciences, San Diego, CA, USA), CD4-APC-Fire750, CD244-PE-D594, CCR7-BV421, HLA-DR-AF700, CD38-BV421, CD28-BV711, CD27-BV650, KLRG-1-APC-Fire750, CD95-PE-CY7 (BioLegend, San Diego, CA, USA), TIGIT-PE-Cy7, LAG-3-APC (Ebioscience, San Diego, CA, USA) and the corresponding isotype controls. Data were acquired with the LSR Fortessa flow cytometer (BD Biosciences) and analyzed with FlowJo software version 10.5 (Tree Star, Ashland, OR, USA). More information about antibodies is listed in the Supplementary Material (Table S2).
Wang X., Wang D., Du J., Wei Y., Song R., Wang B., Qiu S., Li B., Zhang L., Zeng Y., Zhao H, & Kong Y. (2022). High Levels of CD244 Rather Than CD160 Associate With CD8+ T-Cell Aging. Frontiers in Immunology, 13, 853522.
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10

Purification of Interferon-Stimulated PBMCs

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Previously frozen PBMCs from only before and 24 hours after IFN-α2b (41 (link)) were thawed, washed, and incubated with CD3-FITC (BioLegend), CD4-PE/Cy7 (BioLegend), CD8-APC (BD Biosciences), HLA-DR–PE (BioLegend), and CD38-BV421 (BioLegend) antibodies for 40 min at 4°C as per the manufacturers’ recommendation. Immediately before sorting, plasma membrane–compromised cells were labeled with propidium iodide (Sigma-Aldrich). FACS was performed on a MoFlo Legacy Sorter (Beckman Coulter) at the Johns Hopkins School of Public Health Flow Cytometry Core Facility. The population of interest was sorted directly into ≥4 volumes of Quick-RNA MicroPrep lysis buffer (Zymo Research) as per the manufacturer’s recommendation. Sorting was stopped when the number of sorted cells reached 125,000 cells, although many samples did not reach this number. Flow cytometry analysis on two randomly selected post-sort samples revealed >95% purity. Sorted samples were vortexed, incubated for 10 min at room temperature, vortexed again, and frozen at −80°C until isolation.
El-Diwany R., Soliman M., Sugawara S., Breitwieser F., Skaist A., Coggiano C., Sangal N., Chattergoon M., Bailey J.R., Siliciano R.F., Blankson J.N., Ray S.C., Wheelan S.J., Thomas D.L, & Balagopal A. (2018). CMPK2 and BCL-G are associated with type 1 interferon–induced HIV restriction in humans. Science Advances, 4(8), eaat0843.
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