Magnetic bead separation kit

Manufactured by Miltenyi Biotec

Sourced in United States, Germany

The Magnetic bead separation kit is a laboratory equipment used for the isolation and purification of target cells, proteins, or molecules from complex biological samples. The kit utilizes magnetic beads coated with specific ligands or antibodies that bind to the desired target. The magnetic properties of the beads allow for their efficient separation and isolation from the sample using a magnetic field.

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Lab products found in correlation

Ack lysing buffer, by Lonza (3 mentions) Hla dr ab clone l243, by Novus Biologicals (3 mentions) Ack lysing buffer, by Quality Biological (2 mentions) Cd45ra ab negative, by Thermo Fisher Scientific (2 mentions) Phenol red, by Lonza (1 mentions) Gentamicin, by Lonza (1 mentions) Fortessa x20, by BD (1 mentions) Anti cd4 clone gk1, by BioXCell (1 mentions) T cell expansion kit, by Miltenyi Biotec (1 mentions) Phytohemagglutinin (pha), by Merck Group (1 mentions) Anti cd3 ab, by Thermo Fisher Scientific (1 mentions) Anti cd28 ab, by Thermo Fisher Scientific (1 mentions) Serum freex vivo 15 medium, by Lonza (1 mentions) Polymorphprep density gradient, by Axis-Shield (1 mentions) Human ifn γ, by Thermo Fisher Scientific (1 mentions) Roswell park memorial institute (rpmi), by Thermo Fisher Scientific (1 mentions)

Spelling variants of this product

Magnetic beads (547 citations) Magnetic bead separation (70 citations) Magnetic separation (67 citations) Magnetic separation kit (11 citations) Magnetic separation beads (2 citations)

15 protocols using magnetic bead separation kit

Isolation of CCR2+ Hematopoietic Stem Cells

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Example 1

The chemokine receptor CCR2 is expressed on monocyte precursor cells and is required for their entry into the CNS, typically as a chemotactic response to CCL2 which is expressed by KR158B glioma (Clarkson et al., 2015; Sagar et al., 2012; Flores et al., 2015). In this study, it was observed that a subset of HSCs express CCR2 (CCR2 HSC) migrate to intracranial tumor within 3 hours (data not shown). To isolate CCR2⁺ HSCs, bone marrow was collected from non-tumor bearing mice and lineage depleted using magnetic bead separation kit (Miltenyi Biotec, Calif.). The resulting lineage negative HSCs were then further enriched with a secondary magnetic bead depletion using biotinylated anti-CCR2 antibody followed with anti-biotin bead conjugated antibody.

US11464805B2. CCR2+ hematopoietic stem cells mediate T cell activation in adoptive cell therapy (2022-10-11). University of Florida Research Foundation, Incorporated [US]. Inventors: Duane Mitchell [US], Catherine Flores [US].

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Adoptive Transfer of CD8+ T Cells

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Splenocytes were collected from CD45.2 Thy1.1 IFN-γ reporter mice and labeled with 1.25 μM of Carboxyfluorescein succinimidyl ester (CFSE) for 10 minutes at room temperature. The reaction was quenched by the addition of newborn calf serum and cells were washed by centrifugation three times. Cell suspension was then transferred intravenously into CD45.1 recipients that were immediately infected with L. major. For experiments with RAG mice, splenocytes from C57BL/6 mice were collected, red blood cells lysed with ACK lysing buffer (LONZA) and CD8+ T cells were purified using a magnetic bead separation kit (Miltenyi Biotec). Three million CD8+ T cells were transferred into RAG mice that were subsequently infected with L. braziliensis. Mice reconstituted with CD8+ T cells received 4 injections of 250 μg of anti-CD4 within the first 2 weeks in order to ensure that no CD4+ T cells were present.

Novais F.O., Wong A.C., Villareal§ D.O., Beiting D.P, & Scott* P. (2018). CD8+ T cells lack local signals to produce IFN-γ in the skin during leishmania infection. Journal of immunology (Baltimore, Md. : 1950), 200(5), 1737-1745.

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Adoptive Transfer and CD4 Depletion in Leishmaniasis

Cited 1 time

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Spleen cells from C57BL/6 mice were collected, erythrocytes lysed with ACK lysing buffer (Quality Biological, Cat #118-156-101), and CD8⁺ T cells were purified using a magnetic bead separation kit (Miltenyi Biotec, Cat #130-104-075). Rag1^−/− mice were reconstituted with CD8⁺ T cells by intravenous route (3 × 10⁶ cells/mouse) and subsequently were infected with L. braziliensis. Mice received 250 mg of antibody anti-CD4, clone GK1.5 (BioXCell, Cat #BE0003-1, RRID:AB_1107636) by intraperitoneal injections twice a week in the first two weeks post-infection[8 (link)].

Sacramento L.A., Amorim C.F., Lombana C.G., Beiting D., Novais F., Carvalho L.P., Carvalho E.M, & Scott P. (2023). CCR5 promotes the migration of CD8+ T cells to the leishmanial lesions. bioRxiv.

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Isolation and Differentiation of CD14+ Cells

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Autologous CD14⁺ cells were sorted from freshly thawed PBMCs using a magnetic bead-separation kit (Miltenyi Biotec, 130-050-201) according to the manufacturer’s instructions. Isolated CD14⁺ cells were used as targets in the IFN-γ ELISPOT assay directly after sorting or differentiated in vitro into tumor-associated macrophages by culturing with 1 ml fresh X-VIVO 15 medium with Gentamicin and Phenol Red (Lonza, BE02-060Q) and 5% heat-inactivated Human AB Serum, supplemented with 1 ml autologous TCM in 24-well plates for 2 d.

Kjeldsen J.W., Lorentzen C.L., Martinenaite E., Ellebaek E., Donia M., Holmstroem R.B., Klausen T.W., Madsen C.O., Ahmed S.M., Weis-Banke S.E., Holmström M.O., Hendel H.W., Ehrnrooth E., Zocca M.B., Pedersen A.W., Andersen M.H, & Svane I.M. (2021). A phase 1/2 trial of an immune-modulatory vaccine against IDO/PD-L1 in combination with nivolumab in metastatic melanoma. Nature Medicine, 27(12), 2212-2223.

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CD8+ T Cell Adoptive Transfer in Leishmaniasis

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Splenocytes from C57BL/6 WT or perforin-/-, mice were collected, red blood cells lysed with ACK lysing buffer (LONZA) and CD8+ T cells were purified using a magnetic bead separation kit (Miltenyi Biotec). Three million CD8+ T cells were transferred into RAG mice that were subsequently infected with L. braziliensis. Mice reconstituted with CD8+ T cells received 4 injections of 250 μg of anti-CD4 within the first 2 weeks.

Novais F.O., Carvalho A.M., Clark M.L., Carvalho L.P., Beiting D.P., Brodsky I.E., Carvalho E.M, & Scott P. (2017). CD8+ T cell cytotoxicity mediates pathology in the skin by inflammasome activation and IL-1β production. PLoS Pathogens, 13(2), e1006196.

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Monocyte Tracking in Adipose Tissue

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The approach is based on previously published method (26 (link)) with some modifications. Monocytes were isolated from the bone marrow progenitors of WT and α_D-deficient mice using magnetic bead separation kit (Miltenyi Biotec, Gaithersburg, MD, United States). Monocytes were labeled with red, PKH26 (WT) or green, PKH67 (

α_{D}^{- / -}

) fluorescent dyes. Red (1.5 × 10⁶) and green (1.5 × 10⁶) cells were mixed together and injected in tail vein of wild type C57BL6 mice fed high fat diet (45% kcal/fat) for 8 weeks. After 3 days adipose tissue was isolated, digested as described previously (26 (link)) and analyzed using FACS (Fortessa X-20, BD, United States) and imaging flow cytometry (ImageStream Mark II, Amnis).

Cui K., Ardell C.L., Podolnikova N.P, & Yakubenko V.P. (2018). Distinct Migratory Properties of M1, M2, and Resident Macrophages Are Regulated by αDβ2 and αMβ2 Integrin-Mediated Adhesion. Frontiers in Immunology, 9, 2650.

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CD8+ T Cell Adoptive Transfer in Leishmaniasis

Cited 1 time

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Spleen cells from C57BL/6 mice were collected, erythrocytes lysed with ACK lysing buffer (Quality Biological, Cat #118-156-101), and CD8⁺ T cells were purified using a magnetic bead separation kit (Miltenyi Biotec, Cat #130-104-075). Rag1^-/- mice were reconstituted with CD8⁺ T cells by intravenous route (3 x 10⁶ cells/mouse) and subsequently were infected with L. braziliensis. Mice received 250 mg of antibody anti-CD4, clone GK1.5 (BioXCell, Cat #BE0003-1) by intraperitoneal injections twice a week in the first two weeks post-infection [8 (link)].

Amorim Sacramento L., Farias Amorim C., G. Lombana C., Beiting D., Novais F., P. Carvalho L., M. Carvalho E, & Scott P. (2024). CCR5 promotes the migration of pathological CD8+ T cells to the leishmanial lesions. PLOS Pathogens, 20(5), e1012211.

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Murine T-cell Suppression Assay

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T_reg cell suppression assay was performed as previously described (30 (link)). Briefly, naive CD4⁺ T cells from spleen and lymph nodes of C57BL/6 mice were magnetically sorted (Miltenyi) and labeled with CellTrace Violet (CTV) at a concentration of 5 µM according to the manufacturer’s instructions. CD4⁺CD25⁺ T_reg cells were isolated from spleens and lymph nodes of hTNFKI × hTNFR2KI and hTNFKI × hTNFR2^ΔTregs mice using a magnetic bead separation kit (Miltenyi). CTV-labeled T cells were cocultured with T_reg cells in complete RPMI 1640 medium in the presence of 1 µg/mL anti-CD3 (145-2C11; in-house/DRFZ) and 6 ng/mL of anti-CD28 (37.51; in-house/DRFZ) in a humidified atmosphere for 80 h at 37 °C and 5% CO₂. T cell proliferation was analyzed by flow cytometry by measuring the CTV label.

Atretkhany K.S., Mufazalov I.A., Dunst J., Kuchmiy A., Gogoleva V.S., Andruszewski D., Drutskaya M.S., Faustman D.L., Schwabenland M., Prinz M., Kruglov A.A., Waisman A, & Nedospasov S.A. (2018). Intrinsic TNFR2 signaling in T regulatory cells provides protection in CNS autoimmunity. Proceedings of the National Academy of Sciences of the United States of America, 115(51), 13051-13056.

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Adoptive Transfer of CD8 T Cells

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Splenocytes from C57BL/6 mice were collected, red blood cells lysed with ACK lysing buffer (LONZA) and CD8 T cells were purified using a magnetic bead separation kit (Miltenyi Biotec). Three million CD8 T cells were transferred into RAG^−/− mice that were subsequently infected with L. braziliensis. Mice reconstituted with CD8 T cells received 4 injections of 250 μg of anti-CD4 within the first 2 weeks.

Novais F.O., Nguyen B.T, & Scott P. (2020). Granzyme B inhibition by tofacitinib blocks pathology induced by CD8 T cells in cutaneous leishmaniasis. The Journal of investigative dermatology, 141(3), 575-585.

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Isolation and Activation of CD4+ T Cells for mTORC1 Targeting

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CD4⁺ T cells were isolated from PBMCs using a magnetic bead separation kit (Miltenyi Biotec, Bergisch Gladbach, Germany), as previously described in our studies.³³, ³⁵ Briefly, lineage‐specific biotin‐conjugated antibodies and anti‐biotin MicroBeads were used to remove non‐CD4⁺ T cells, and CD25⁺ PE‐labelled cells were magnetically removed from purified CD4⁺ fractions, leaving unlabeled CD4⁺CD25⁻ cells for stimulation experiments. The CD4⁺ T cells were activated for 72 h in RPMI‐1640 medium supplemented with 10% FBS and a T Cell Expansion Kit (Miltenyi Biotec), containing biotinylated antibodies against human CD2, CD3 and CD28. For pathogen stimulation experiments, cells were stimulated with phytohemagglutinin (PHA, 5 μg mL⁻¹, Sigma‐Aldrich, Missouri, USA) for 72 h, followed by a further sorting analysis. To target mTORC1, commercial non‐targeted siRNA, PTEN siRNA and PS6K siRNA were used following mitogenic stimulation according to the manufacturer's instructions (#6568, #6251 and #6566, Cell Signalling, Danvers, USA).

Chen H., Liu Z., Zha J., Zeng L., Tang R., Tang C., Cai J., Tan C., Liu H., Dong Z, & Chen G. (2023). Glucocorticoid regulation of the mTORC1 pathway modulates CD4+ T cell responses during infection. Clinical & Translational Immunology, 12(8), e1464.

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