Cd19 bv785

Manufactured by BioLegend

CD19-BV785 is a fluorescent-conjugated antibody that binds to the CD19 antigen. CD19 is a transmembrane protein expressed on the surface of B cells and is commonly used as a marker for identifying and analyzing B cell populations.

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5 protocols using cd19 bv785

Bone Marrow Transplantation and Hematopoietic Reconstitution

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CD45.1 Ahr^+/+ (B6.SJL-Ptprc^a/BoyAiTac; 4007 F), CD45.2 Ahr^+/+ (C57BL/6NTac; B6-F), and CD45.2 Ahr^-/- mice were lethally irradiated (2 × 6 Gy, 6 hr apart) at 5–7 weeks of age. Bone marrow from donor CD45.1 Ahr^+/+ and CD45.2 Ahr^-/- mice was delivered by tail vein injection 1 hr after the second radiation dose. Mice were maintained for 2 weeks on autoclaved food and water containing 2 mg/ml neomycin sulfate (VWR 89149–866) and 1000 U/ml polymyxin B (Millipore Sigma P4932-5MU). Bone marrow engraftment was assessed 8 weeks after transplantation by processing 10 μl of tail vein blood as described above and staining with Live/Dead Fixable Blue stain (Fisher L34962) and the following antibodies: CD45.2 PerCP-Cy5.5 (Fisher/Invitrogen 45-0454-80), NK-1.1 FITC (Biolegend 108706), CD11c PE-Cy7 (Fisher/Invitrogen 25-0114-82), CD45.1 PE (Biolegend 110707), CD19 BV 785 (Biolegend 115543), CD3 BV 650 (Biolegend 100229), CD8a BV 510 (Biolegend). Mice were infected 9 weeks after transplantation. Sampling was performed as described in Longitudinal infection monitoring with the following modification: 12 μl of blood for BUN and heme quantification was collected on day 0, and 7–9 only. Flow panel used for validation.

Lissner M.M., Cumnock K., Davis N.M., Vilches-Moure J.G., Basak P., Navarrete D.J., Allen J.A, & Schneider D. (2020). Metabolic profiling during malaria reveals the role of the aryl hydrocarbon receptor in regulating kidney injury. eLife, 9, e60165.

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Synthesis and Characterization of OSU-2S

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OSU-2S was synthesized as described [7 (link)] at The Ohio State University Medicinal Chemistry shared resource and verified by Nuclear Magnetic Resonance and Mass Spectrometry. DMSO (Invitrogen) was used to dissolve OSU-2S for in-vitro studies. Antibodies used include c-Myc, p21 and β-Actin (Cell Signaling, Danvers, MA), CD45-FITC, CD3-PE-Cy7, CD19-BV785 (BioLegend, San Diego, CA), murine CD45-PECF594 (BD Biosciences) and ROR1- PE (R&D Systems).

Goswami S., Chiang C.L., Zapolnik K., Nunes J., Ventura A., Mo X., Xie Z., Lee L.J., Baskar S., Rader C., Byrd J.C., Phelps M., Bhatnagar B, & Muthusamy N. (2022). ROR1 targeted immunoliposomal delivery of OSU-2S shows selective cytotoxicity in t(1;19)(q23; p13) translocated B-cell acute lymphoblastic leukemia. Leukemia research, 118, 106872.

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Characterizing Immune Cell Activation

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Fresh PBMCs were cultured either alone, with Pam₃CysSK₄ or XS15, or a mix of phytohaemagglutinin-L (PHA) and Pokeweed mitogen (PWM). Adherent and non-adherent cells were stained with mAbs: CD3-BV711, CD56-BV421, CD19-BV785, Zombie aqua (all Biolegend), CD14-Alexa Fluor 700, HLA-DR-PerCP, and CD69-APC-Cy7 (all BD). Cells were measured by flow cytometry as described above.

Rammensee H.G., Wiesmüller K.H., Chandran P.A., Zelba H., Rusch E., Gouttefangeas C., Kowalewski D.J., Di Marco M., Haen S.P., Walz J.S., Gloria Y.C., Bödder J., Schertel J.M., Tunger A., Müller L., Kießler M., Wehner R., Schmitz M., Jakobi M., Schneiderhan-Marra N., Klein R., Laske K., Artzner K., Backert L., Schuster H., Schwenck J., Weber A.N., Pichler B.J., Kneilling M., la Fougère C., Forchhammer S., Metzler G., Bauer J., Weide B., Schippert W., Stevanović S, & Löffler M.W. (2019). A new synthetic toll-like receptor 1/2 ligand is an efficient adjuvant for peptide vaccination in a human volunteer. Journal for Immunotherapy of Cancer, 7, 307.

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Multicolor Flow Cytometry Panel

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The following fluorochrome conjugated anti-human antibodies were used: CD3 APC efluor 780 (eBioscience #47-0036-42) or BV650 (Biolegend #317324), CD4 BV 510 (BD Horizon/BD Biosciences #562970), CD8 evolve 655 (eBioscience #86-0088-42), CD33 APC (eBioscience#17-0338-42), CD56 efluor 710 (eBioscience #46-056-42) or FITC (Biolegend #304604), PD-1 BV 785 (BioLegend #329930), CCR7 BV785 (Biolegend #353230), CD19 BV785 (Biolegend #302240), CD62L BV785 (Biolegend #304830), IFN-γ efluor 450 (eBioscience #48-7319-42), Perforin Alexa Fluor 647 (Biolegend #353322) or PE (Biolegend #353304), granzyme FITC (Biolegend #515403). Appropriate isotype control antibodies were employed to determine the specificity of test antibodies.

Vardam-Kaur T., Pathangey L.B., McCormick D.J., Bergsagel P.L., Cohen P.A, & Gendler S.J. (2021). Multipeptide stimulated PBMCs generate TEM/TCM for adoptive cell therapy in multiple myeloma. Oncotarget, 12(20), 2051-2067.

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Comprehensive Immune Profiling in Mice

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Mice were anesthetized with 1.5–2% isoflurane and blood was collected using cardiac puncture. Organs were perfused with cold PBS and spleen was harvested and weighed. Flow cytometry was performed as described previously (20 (link), 25 (link)). Following antibodies (with their respective clone numbers) were obtained from eBioscience unless otherwise stated: 7-AAD, MHCII-FITC (M5/114.15.2), Gr-1-APC (1A8), Ly6C-eF450 (HK1.4), CD11b-SuperBright600 (M1/70), CD45.2-BV650 (104; BioLegend), F4/80-APC-eF780 (BM8), CD11c-BV785 (N418; BioLegend), NK1.1-PE (PK136), CD3e-APC (145-2C11), CD8a-eF450 (53-6.7), CD4-SuperBright600 (RM4-5), CD19-BV785 (6D5; BioLegend). AccuCheck beads (Life Technologies) were used to determine absolute numbers by normalizing to tissue mass. Data were collected with a Becton-Dickinson LSRII analyzer and analyzed using FlowJo software (Trecstar Software).

Zarjou A., Black L.M., McCullough K.R., Hull T.D., Esman S.K., Boddu R., Varambally S., Chandrashekar D.S., Feng W., Arosio P., Poli M., Balla J, & Bolisetty S. (2019). Ferritin Light Chain Confers Protection Against Sepsis-Induced Inflammation and Organ Injury. Frontiers in Immunology, 10, 131.

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