Mojosort mouse pan b cell isolation kit 2

Manufactured by BioLegend

Sourced in United States

The MojoSort™ Mouse Pan B Cell Isolation Kit II is a laboratory tool designed for the isolation of pan B cells from mouse samples. It provides a simple and efficient method to separate B cells from other cell types present in the sample.

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Lab products found in correlation

S3 sorter, by Bio-Rad (1 mentions) Anti mouse cd16 cd32 mab, by BioLegend (1 mentions) Rneasy micro kit, by Qiagen (1 mentions) Rneasy kit, by Qiagen (1 mentions) Cobalt chloride, by Merck Group (1 mentions) Lymphoprep, by STEMCELL (1 mentions) Rpmi 1640 medium, by Thermo Fisher Scientific (1 mentions) Mojosort mouse cd3 t cell isolation kit, by BioLegend (1 mentions) Corn oil, by Merck Group (1 mentions) Tam diet, by Inotiv (1 mentions) Tamoxifen, by Thermo Fisher Scientific (1 mentions) Tamoxifen, by Inotiv (1 mentions)

5 protocols using mojosort mouse pan b cell isolation kit 2

Adoptive B cell transfer in helminth infection

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Cells were isolated from mes-LN and BM on day 28 or later after infection and were purified with the MojoSort™ Mouse Pan B Cell Isolation Kit II (BioLegend). 2-3x10⁶ cells were transferred i.v. into Ly5.1_IgH^a recipient mice which had been infected with N. brasiliensis three days before. Mice were analyzed on day 10 after infection. BM, mes-LN and med-LN samples were taken for flow cytometric analysis and serum was used for IgE ELISA.

Haase P., Schäfer S., Gerlach R.G., Winkler T.H, & Voehringer D. (2022). B cell fate mapping reveals their contribution to the memory immune response against helminths. Frontiers in Immunology, 13, 1016142.

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Isolation and Sequencing of B Cells from Infected Mice

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Cells were isolated from mes-LN and BM on day 28 or later after infection and on day 10 after transfer and B cells were purified with the MojoSort™ Mouse Pan B Cell Isolation Kit II (BioLegend). For cells after transfer Fc receptors were blocked with anti-mouse CD16/CD32 mAb and the cells stained at 4°C for 30 min for CD45.2 (Biolegend). Cells were sorted for tdTomato⁺ and tdTomato^- before transfer and CD45.2⁺ tdTomato⁺ or CD45.2⁺ Tomato^- cells after transfer using the S3 sorter (Bio-Rad). The cells were sorted into 1 ml FCS, pelleted and immediately lysed with RLT-buffer (RNeasy-Kit; Qiagen). The lysate was stored at -80°C until RNA isolation. RNA was isolated using the RNeasy micro kit (Qiagen). The samples were prepared for sequencing and analyzed according to the procedures described in (31 (link)).Samples were sequenced in an illumina MiSeq machine using the MiSeq Reagent Kit v3 (600-cycle).

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Hypoxia and AhR Activation in B Cells

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B cells were purified from the spleen by negative selection using the MojoSort™ Mouse Pan B Cell Isolation Kit II (Biolegend, San Diego, CA, USA) following the manufacturer’s instructions. The isolated B cell population contained at least 90% of CD19+CD5+-double positive cells. B cells were cultured in RPMI supplemented with 10% FBS, 50 µM 2-β-mercaptoethanol, 100 U/mL penicillin, and 100 µg/mL streptomycin. To mimic hypoxia, 150 µM cobalt chloride (Sigma-Aldrich, Burlington, MA, USA) was added, and the cells were cultured overnight at 37 °C and 5% CO₂. In order to activate AHR, the cells were incubated with 250 nM FICZ (Sigma-Aldrich, Burlington, MA, USA) for 2 hours at 37 °C and 5% CO_2.

Gonder S., Largeot A., Gargiulo E., Pierson S., Fernandez Botana I., Pagano G., Paggetti J, & Moussay E. (2021). The Tumor Microenvironment-Dependent Transcription Factors AHR and HIF-1α Are Dispensable for Leukemogenesis in the Eµ-TCL1 Mouse Model of Chronic Lymphocytic Leukemia. Cancers, 13(18), 4518.

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Isolation and Expansion of Primary T and B Cells

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Peripheral blood mononuclear cells were harvested from blood donor buffy coats via density gradient isolation. Buffy coats were carefully pipetted onto STEMCELL Technologies Lymphoprep and centrifuged at 600 x g for 30 min. The resulting mononuclear cell layer was extracted, washed 4 times in PBS and up to 1e9 cells put into culture in ThermoFisher RPMI 1640 medium with 10% FCS, Pen/Strep and HEPES buffer. T cells were stimulated with 1000 U mL⁻¹ IL-2, 200 ng mL⁻¹ OKT3 and 50 ng mL⁻¹ 15E8 for two days before transduction.
Primary mouse cells were gathered by singularizing the spleen of C57BL/6J (Black 6; RRID:IMSR_JAX:000664) mice and isolating either B cells with a MojoSort Mouse Pan B Cell Isolation Kit II (BioLegend Cat#480087) or T cells with a MojoSort Mouse CD3 T cell Isolation Kit (BioLegend Cat#480031) according to the manufacturer’s instructions. Murine B cells were cultured in RPMI 1640 supplemented with 10% FCS and Pen/Strep, murine T cells were cultured in X-Vivo 15 supplemented with 5% FCS and stimulated with 200 ng mL⁻¹ anti-CD3, 100 ng mL⁻¹, 1000U mL⁻¹ IL-2 and 10 ng mL⁻¹ IL-15 for three days prior to transduction.

Prinz L.F., Riet T., Neureuther D.F., Lennartz S., Chrobok D., Hübbe H., Uhl G., Riet N., Hofmann P., Hösel M., Simon A.G., Tetenborg L., Segbers P., Shimono J., Gödel P., Balke-Want H., Flümann R., Knittel G., Reinhardt H.C., Scheid C., Büttner R., Chapuy B., Ullrich R.T., Hallek M, & Chmielewski M.M. (2024). An anti-CD19/CTLA-4 switch improves efficacy and selectivity of CAR T cells targeting CD80/86-upregulated DLBCL. Cell Reports Medicine, 5(2), 101421.

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Adoptive Transfer of Memory B Cells

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Tamoxifen (Fisher) was dissolved in Corn Oil (Sigma–Aldrich) at 20 mg/m and injected at 2 mg per 20 g mouse i.p. Mice were administered two doses of Tamoxifen on days 4 and 6 p.i., unless otherwise noted. TAM diet (Envigo) containing chow replaced normal chow on the day after the final Tamoxifen dose. To overcome initial taste aversion, additional crushed TAM diet was placed in the cage at the time of the first TAM diet feeding. For adoptive transfer experiments, B cells were enriched using a MojoSort Mouse Pan B Cell Isolation Kit II (480088) (Biolegend). Depletion of CD62L⁺ B cells was achieved via addition of Biotin anti-mouse CD62L (5011606) (eBioscience). The number of S1pr2-Tomato⁺ MBCs within the enriched CD62L⁻ B cell population was determined using flow cytometry. CD62L⁻ B cells were then transferred into recipient mice such that each recipient received 10–30,000 S1pr2-Tomato⁺ MBCs. For the recall experiments, CD62L⁻ and CD62L⁺S1pr2-Tomato⁺ cells were sorted from the enriched B cell population. The number of MBCs within each sorted population was quantified by flow cytometry. Sorted CD62L⁻ and CD62L⁺S1pr2-Tomato⁺ B cells were then transferred into recipient mice such that each recipient received 15–20,000 MBCs.

Hanson C.H., Henry B., Andhare P., Lin F.J., Pak H., Turner J.S., Adams L.J., Liu T., Fremont D.H., Ellebedy A.H, & Laidlaw B.J. (2023). CD62L expression marks a functionally distinct subset of memory B cells. Cell reports, 42(12), 113542.

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