Cd14 apc clone m5e2

Manufactured by BD

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CD14-APC (clone M5E2) is a flow cytometry reagent that binds to the CD14 surface antigen. CD14 is a glycosylphosphatidylinositol (GPI)-anchored protein expressed primarily on the surface of monocytes and macrophages. The CD14-APC conjugate allows for the identification and enumeration of CD14-positive cells in flow cytometric analysis.

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CD14-APC (49 citations) CD14 (clone M5E2, (21 citations) CD14 (M5E2, (13 citations) APC clone M5E2 (3 citations) CD14-allophycocyanin (APC; clone M5E2), (2 citations) Anti-CD14 APC (clone M5E2, (2 citations)

5 protocols using cd14 apc clone m5e2

Comprehensive PBMC Immunophenotyping Protocol

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The frozen PBMC samples were thawed, the number of total cells was counted (Beckman Coulter AcT/Diff). Next, relative fractions of T cells (CD3+/CD4+ & CD3+/CD8+), B cells (CD19+/CD20+), monocytes (CD14+) and NK cells (CD16+/CD56+) were determined in freshly thawed and unstimulated PBMCs via flow cytometry using standard protocols for FACS surface staining. Briefly, cells were stained with following antibodies using two panels: CD3‐Alexa Fluor 488 (clone UCHT1), CD4‐PerCP‐Cy5.5 (clone RPA‐T4), CD8‐APC‐H7 (clone SK1), CD19‐PE‐Cy7 (clone SJ25C1), CD20‐Alexa Fluor 700 (2H7), CD16‐PE (clone 3G8), CD56‐PE (clone MY31) and CD14‐APC (clone M5E2, all antibodies were purchased from BD Pharmingen). Additionally, both panels were stained with CD45‐PE‐TexasRed (clone HI30, Invitrogen). In addition, cell viability was checked and percentage of dead cells was determined. Cells were analysed using a BD FACS Canto II flow cytometer and data was analysed using the FlowJo software (BD Biosciences).

Wisgrill L., Fyhrquist N., Ndika J., Paalanen L., Berger A., Laatikainen T., Karisola P., Haahtela T, & Alenius H. (2022). Bet v 1 triggers antiviral‐type immune signalling in birch‐pollen‐allergic individuals. Clinical and Experimental Allergy, 52(8), 929-941.

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Immunophenotypic Analysis of Stem Cells

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Flow cytometric analysis of hematopoietic surface markers was analyzed on a FACS Canto II (BD Biosciences, Franklin Lakes, New Jersey, USA) with the following antibodies: CD3-PerCP-Cy5.5 (clone UCHT1), CD19-FITC (clone HIB19), CD31-PE (clone WM59), CD34-APC (clone 581), CD38-PE (clone HIT2), CD43-FITC (clone 1G10), CD14-PE (clone MOP9), CD66b-PE (clone G10F5) (all BD Bioscience), CD45-APC-Cy7 (clone 2D1), CD117-PE-Cy7 (clone 104D2), CD11c-PE-Cy7 (clone 3.9), HLA-DR-FITC (clone LN3), CD235a-Pacific Blue (clone 6A7M) (all eBioscience, San Diego, California, USA), CD33-APC (clone AC104.3E3; Miltenyi Biotec). EBs were dissociated with Accutase (Stemcell Technologies, Vancouver, British Columbia, Canada) for 10–15 min prior to staining with antibodies. Single cells were incubated with 1% human IgG solution (Privigen, CSL Behring, King of Prussia, Pennsylvania, USA) for 30 min at 4 °C to block unspecific binding.
Immunophenotypic analysis of MSCs and iMSCs was performed with CD14-APC (clone M5E2), CD29-PE (clone MAR4), CD31-PE (clone WM59), CD34-APC (clone 581), CD45-APC (clone HI30), CD73-PE (clone AD2), CD90-APC (clone 5E10) (all BD Biosciences), and CD105-FITC (clone MEM-226; ImmunoTools, Friesoythe, Germany). Data was analyzed using the FlowJo software (Tree Star, Ashland, Oregon, USA).

Cypris O., Frobel J., Rai S., Franzen J., Sontag S., Goetzke R., Szymanski de Toledo M.A., Zenke M, & Wagner W. (2019). Tracking of epigenetic changes during hematopoietic differentiation of induced pluripotent stem cells. Clinical Epigenetics, 11, 19.

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Flow Cytometry Immunophenotyping Protocol

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The following antibodies were used: CD34-PE (clone 563, BD Pharmingen, RRID:AB_393871), CD45-APC (clone HI30, BD Pharmingen, RRID:AB_398600), CD14-APC (clone M5E2, BD Pharmingen, RRID:AB_398596), CD15-BV785 (clone W6D3, BioLegend, RRID:AB_2632921), CD16-BV510 (clone 3G8, BD Horizon, RRID:AB_2744296). Cell viability was assessed with DAPI (Life Technologies). Cells were assayed on a BD Fortessa and data were analyzed with FlowJo software (Tree Star, RRID:SCR_008520).

Wheeler E.C., Vora S., Mayer D., Kotini A.G., Olszewska M., Park S.S., Guccione E., Teruya-Feldstein J., Silverman L., Sunahara R.K., Yeo G.W, & Papapetrou E.P. (2022). Integrative RNA-omics discovers GNAS alternative splicing as a phenotypic driver of splicing factor-mutant neoplasms. Cancer discovery, 12(3), 836-855.

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Immunophenotyping of Mesenchymal Stromal Cells

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Immunophenotype of MSCs and iMSCs was analyzed with a fluorescence-activated cell sorter (FACS) Canto II (BD Biosciences, Franklin Lakes, USA) after staining with the following antibodies, as described before [23 (link)]: CD29 phycoerythrin (PE; clone MAR4; BD), CD34 allophycocyanin (APC; clone 581; BD), CD31 PE (clone WM59; BD), CD45 APC (clone HI30; BD), CD105 fluorescein isothiocyanate (FITC; clone MEM-226; ImmunoTools, Friesoythe, Germany), CD90 APC (clone 5E10; BD), CD73 PE (clone AD2; BD), and CD14 APC (clone M5E2; BD).

Hollmann J., Brecht J., Goetzke R., Franzen J., Selich A., Schmidt M., Eipel M., Ostrowska A., Hapala J., Fernandez-Rebollo E., Müller-Newen G., Rothe M., Eggermann T., Zenke M, & Wagner W. (2020). Genetic barcoding reveals clonal dominance in iPSC-derived mesenchymal stromal cells. Stem Cell Research & Therapy, 11, 105.

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Flow Cytometry Antibody Staining

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For flow cytometry, the following antibodies were used: CD34-PE
(clone 563, BD Pharmingen), CD41a- PE-Cy7 (clone HIP8, BD Pharmingen),
CD90-PerCP/Cy5.5 (clone 5E10, BioLegend), CD45-APC (clone HI30, BD
Pharmingen), CD15-FITC (clone W6D3, BD Pharmingen), CD14-APC (clone M5E2, BD
Pharmingen), CD33-PE-CF594 (clone WM53, BD Horizon), CD235a-PerCP/Cy5.5
(clone HI264, BioLegend), CD7-BV510 (Clone M-T701, BD Horizon), CD4-PE-Cy7
(Clone RPA-T4, BD Pharmingen) and CD8-PerCP/Cy5.5 (Clone RPA-T8, BD
Pharmingen). Cell viability was assessed with DAPI (Life Technologies).
Cells were then assayed on a BD Fortessa and data were analyzed with FlowJo
software (Tree Star). Sorting of CD41a⁺ and
CD41a⁻ cells was performed on a BD FACS Aria II.

Kotini A.G., Chang C.J., Chow A., Yuan H., Ho T.C., Wang T., Vora S., Solovyov A., Husser C., Olszewska M., Teruya-Feldstein J., Perumal D., Klimek V.M., Spyridonidis A., Rampal R.K., Silverman L., Reddy E.P., Papaemmanuil E., Parekh S., Greenbaum B.D., Leslie C.S., Kharas M.G, & Papapetrou E.P. (2017). Stage-specific human induced pluripotent stem cells map the progression of myeloid transformation to transplantable leukemia. Cell stem cell, 20(3), 315-328.e7.

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