Cd45 j33

Manufactured by Beckman Coulter

Sourced in United States

The CD45 (J33) is a laboratory equipment product from Beckman Coulter. It is a monoclonal antibody used for the identification and enumeration of cells expressing the CD45 antigen, which is present on the surface of leukocytes. The core function of the CD45 (J33) is to provide a tool for the analysis and characterization of cells in flow cytometry applications.

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Lab products found in correlation

Cd105 43a3, by BioLegend (2 mentions) Cd34 8g12, by BD (2 mentions) Af6 120, by Thermo Fisher Scientific (2 mentions) Epcam 9c4, by BioLegend (1 mentions) Cd81 5a6, by BioLegend (1 mentions) Fitc labelled secondary antibody, by Santa Cruz Biotechnology (1 mentions) Mouse igg, by Santa Cruz Biotechnology (1 mentions) Rituximab, by Genentech (1 mentions) Rhil 12, by Thermo Fisher Scientific (1 mentions) Rhil 15, by CellGenix (1 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions) Penicillin, by Thermo Fisher Scientific (1 mentions) Streptomycin, by Thermo Fisher Scientific (1 mentions) Resiquimod r848, by Merck Group (1 mentions) Cd38 ls198 4 3, by Beckman Coulter (1 mentions)

5 protocols using cd45 j33

Multivariate Flow Cytometry and Western Blotting

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To characterise MV flow cytometrically, the following antibodies were used: CD45 (J33, #A07782), CD235a (11E4B-7–6, #A07792, both Beckman Coulter), CD62P (AK4, #304905), CD47 (CC2C6, #323109), MUC1 (16A, #355604) EGFR (AY13, #352904), EMMPRIN (HIM6, #306207), CD62E (HAE-1f, #336008), EpCAM (9C4, #324208, all from Biolegend). To analyse CD63 expression, vesicles were incubated with the unlabelled CD63 antibody (H5C6, #556019, BD) or mouse IgG (#sc-2025, Santa Cruz Biotechnology) and signals visualised using a FITC-labelled secondary antibody (#sc-2010, Santa Cruz Biotechnology). Protein expression by Western Blotting was assessed using primary antibodies against EMMPRIN (H200, #sc-13976), MUC1 (VU4H5, #sc-7313), Tsg101 (C-2, #sc-7964), Hsp90 (F-8, #sc-13119, all from Santa Cruz Biotechnology), HDAC1 (#2062), EGFR, (D38B1, #4267, both from Cell Signaling), CD9 (MEM-61, #21270091, Immunotools), CD81 (5A6, #349501, Biolegend), Wnt5a (442625, #MAB645, R&D systems), syntenin (EPR8102, #ab133267, Abcam), GM130 (35/GM130, #610823, BD Transduction Laboratories) or Tubulin (DM1A, #05–829, Millipore). HRP-labelled secondary antibodies were purchased from Santa Cruz Biotechnology (#sc-2004, #sc-2005, #sc-2006).

Menck K., Bleckmann A., Wachter A., Hennies B., Ries L., Schulz M., Balkenhol M., Pukrop T., Schatlo B., Rost U., Wenzel D., Klemm F, & Binder C. (2017). Characterisation of tumour-derived microvesicles in cancer patients’ blood and correlation with clinical outcome. Journal of Extracellular Vesicles, 6(1), 1340745.

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Isolation and Characterization of Luci-eASCs

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Luci-eASCs were defined according to the criteria of the International Society for Cellular Therapy (43 (link)). Luci-eASCs were characterized by their immunophenotype; positive for CD73 (AD2), CD90 (5E10, both from Becton Dickinson), and CD105 (43A3, from Biolegend) and negative for CD14 (RM052, from Immunotech), CD34 (8G12, from Becton Dickinson), CD45 (J33, from Beckman Coulter), and HLA-DR (AF6-120.1, from eBiosciences).

Lopez-Santalla M., Mancheño-Corvo P., Escolano A., Menta R., DelaRosa O., Abad J.L., Büscher D., Redondo J.M., Bueren J.A., Dalemans W., Lombardo E, & Garin M.I. (2017). Biodistribution and Efficacy of Human Adipose-Derived Mesenchymal Stem Cells Following Intranodal Administration in Experimental Colitis. Frontiers in Immunology, 8, 638.

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Characterization of NK Cell Function

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The following anti-human monoclonal antibodies (mAbs) were used: CD56(N901), CD3(UCHT1), CD159a(Z199), CD158a,h(EB6B), and CD45(J.33; all Beckman Coulter); CD16(3G8), CD158b(CH-L), IFNγ(B27), CD107a(H4A3), Syk(4D10), ZAP-70(1E7.2), pZAP70(pY319)/pSYK(pY352), pERK1/2(pT202/pY204), pAkt(pS473; all BD); CD3z(6B10.2; eBioscience); CD158e1(DX9), CD107a(H4A3), goat anti-mouse IgG(Poly4053; all Biolegend); FcεR1γ (Milli-Mark); and rituximab (Genentech). The following endotoxin-free recombinant human (rh) cytokines were used: rhIL-12, rhIL-18 (Peprotech), rhIL-15 (CellGenix and Miltenyi). The following cell lines were used: K562 (ATCC, CCL-243) and Raji (ATCC, CCL-86). Cells were obtained from ATCC in 2008, viably cryopreserved and stored in LN2, thawed for use in these studies, and maintained for <2 months at a time in continuous culture as per ATCC instructions. K562 cells were authenticated in 2015 by confirming cell growth morphology (lymphoblast), short tandem repeat profile, growth characteristics, and functionally as NK cell sensitive targets. Raji cells were authenticated in 2015 by confirming cell growth morphology (lymphoblast), short tandem repeat profile, growth characteristics, phenotype of uniform expression of human CD20, and functionally as anti-CD20 mAb opsonized targets for ADCC.

Wagner J.A., Berrien-Elliott M.M., Rosario M., Leong J.W., Jewell B.A., Schappe T., Abdel-Latif S, & Fehniger T.A. (2016). Cytokine-induced memory-like differentiation enhances unlicensed NK cell anti-leukemia and FcγRIIIa-triggered responses. Biology of blood and marrow transplantation : journal of the American Society for Blood and Marrow Transplantation, 23(3), 398-404.

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Polyclonal Activation of B Cells

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Polyclonal B–cell activation was carried out by stimulating 2 × 10⁶ PBMC/well in a total volume of 2 mL/well in 24–well plates with an activation cocktail consisting of 2.5 μg/mL Toll–like receptor 7/8 agonist (resiquimod [R848]; Sigma–Aldrich, St. Louis, MO) and 1000 IU/mL IL–2 (Proleukin, Novartis, the Netherlands).^{15 (link)} Peripheral blood mononuclear cell cultures were carried out in Iscove’s modified Dulbecca’s medium (Gibco Invitrogen, Paisley, UK) containing 10% fetal bovine serum (Gibco Invitrogen) and 100 U/mL penicillin with 100 μg/ml streptomycin (Gibco Invitrogen). Supernatants harvested at day 6 (d6) or day 10 (d10) were kept at −20°C until further use. Flow cytometry was performed before (d0) and after (d6, d10, and d14) polyclonal activation according to standard protocols using the following antibodies (clone): CD19 (A07770), CD27 (1A4CD27), CD38 (LS198–4–3) and CD45 (J33) (all from Beckman Coulter, Woerden, the Netherlands).

Karahan G.E., Krop J., Wehmeier C., de Vaal Y.J., Langerak–Langerak J., Roelen D.L., Lardy N.M., Bemelman F.J., ten Berge I.J., Reinders M.E., van Kooten C., Claas F.H, & Heidt S. (2019). An Easy and Sensitive Method to Profile the Antibody Specificities of HLA–specific Memory B Cells. Transplantation, 103(4), 716-723.

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Luci-eASCs Characterization Criteria

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Luci-eASCs were defined according to the criteria of the International Society for Cellular Therapy [36 (link)]. Luci-eASCs were stained using antibodies for CD73 (AD2), CD90 (5E10, both from Becton Dickinson), CD105 (43A3, Biolegend, San Diego, CA, USA), CD14 (RM052, Immunotech, Monrovia, CA, USA), CD34 (8G12, Becton Dickinson, Franklin Lakes, NJ, USA), CD45 (J33, Beckman Coulter, Brea, CA, USA) and HLA-DR (AF6-120.1, eBiosciences, Waltham, MA, USA) [18 (link)].

Lopez-Santalla M., Mancheño-Corvo P., Escolano A., Menta R., Delarosa O., Redondo J.M., Bueren J.A., Dalemans W., Lombardo E, & Garin M.I. (2018). Comparative Analysis between the In Vivo Biodistribution and Therapeutic Efficacy of Adipose-Derived Mesenchymal Stromal Cells Administered Intraperitoneally in Experimental Colitis. International Journal of Molecular Sciences, 19(7), 1853.

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