Cd28 pe

Manufactured by Beckman Coulter

Sourced in Italy, United States

The CD28 PE is a fluorescently labeled antibody that binds to the CD28 receptor on the surface of T cells. It is a commonly used reagent in flow cytometry applications for the identification and quantification of CD28-positive T cell populations.

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4 protocols using cd28 pe

Phenotypic Characterization of Cell Subsets

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Various subpopulations within the cell products (PBMCs prior to culture and cultured DC-CIKs) were identified by flow cytometry, as previously described (11 (link)), using the following fluorochrome-conjugated antibodies: CD3 PerCP-Cy5.5, CD4 FITC, CD8 FITC, CD25 PE, CD28 PE, CD56 PE (all Beckman Coulter, Inc.), PD-1 PE, lymphocyte-activation gene 3 (LAG-3) PE, tumor necrosis factor receptor superfamily member 9 (4-1BB; CD137) PE and T cell immunoglobulin and mucin protein 3 (TIM-3) PeCy-7 (all BioLegend, Inc.). Three-color flow cytometry analysis was performed on a Cytomics FC500 flow cytometer with CXP analysis software (Beckman Coulter, Inc.).

Huang L., Qiao G., Morse M.A., Wang X., Zhou X., Wu J., Hobeika A., Ren J, & Lyerly H.K. (2019). Predictive significance of T cell subset changes during ex vivo generation of adoptive cellular therapy products for the treatment of advanced non-small cell lung cancer. Oncology Letters, 18(6), 5717-5724.

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Quantifying Immune Cell Subsets

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The PBMC were incubated with combinations of fluorescein isothiocyanate (FITC), phycoerythrin (PE), PE-cyanine 5.5 (PE-cy5.5), and peridinin chlorophyll protein monoclonal antibodies. The monoclonal antibodies were CD3-FITC, CD3-Percp/Cy5.5, CD4-PE, CD4-FITC, CD8-FITC, CD8-PE, CD16-FITC, CD56-PE, CD19-PE, CD25-FITC, CD127-Percp/Cy5.5, and CD28-PE (Beckman Coulter). About 10,000 lymphocytes were assessed with FC500 software to determine the percentage of CD3⁺, CD4⁺, CD8⁺, CD3⁻CD16⁺CD56⁺, CD19⁺, CD4⁺CD25⁺CD127⁺, CD8⁺CD28⁻, and CD8⁺CD28⁺ lymphocytes.

Yao J., Guan S., Liu Z., Li X, & Zhou Q. (2020). Changes in immune indicators and bacteriologic profile were associated with patients with ventilator-associated pneumonia. Medicine, 99(16), e19716.

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Multiparametric Flow Cytometry Immunophenotyping

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Freshly EDTA peripheral blood from controls and patients were stained with suitable conjugated antibody: APC-CD8, FITC-CD4, PE-CD28, Pe-Cy-5-CD27, Pe-Cy-5-CD25, PE-CD62L, PE-CD69, Pe-Cy-5-CD14 and PE-CD16 (all Beckmann Coulter, Milan, Italy). Cells were incubated for 15 min at room temperature and for other 10 minute with VersaLyse reagent (Beckmann Coulter) to lyse red blood cells. Data were acquired using a FC500 (Beckmann Coulter) flow cytometer and analyzed using Kaluza software. The area of positivity was determined using an isotype-matched mAb, a total of 10⁴ events for each sample were acquired.

Faienza M.F., Ventura A., Piacente L., Ciccarelli M., Gigante M., Gesualdo L., Colucci S., Cavallo L., Grano M, & Brunetti G. (2014). Osteoclastogenic Potential of Peripheral Blood Mononuclear Cells in Cleidocranial Dysplasia. International Journal of Medical Sciences, 11(4), 356-364.

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T-cell Subset and Senescence Profiling

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Blood samples were analyzed centrally at the Bordeaux University Hospital Laboratory of Immunology by flow cytometry using a NAVIOS flow cytometer from Beckman-Coulter (France). All EDTA anticoagulated blood samples were analyzed fresh within a working day after blood withdrawal. A panel of 9-color markers was used with the following antibodies: FITC-CD57, PE-CD28, PC5.5-CD38, ECD-CD45RA, PC7-CD27, APC-CD4, APC Alexa 750-CD3, and APC Alexa 700-CD8 (Beckman-Coulter); V450-HLA-DR (Becton Dickinson, New Jersey, USA). Events were acquired with the CXP software and analyzed on Kaluza software. The phenotypic marker CD27 was used with CD45RA to divide CD4 + and CD8 + T cells into four different subsets: CD45RA + CD27 + [naive T cells (TN)]; CD45RA -CD27 + [central memory T cells (TCM)]; CD45RA -CD27 - [effector memory T cells (TEM)]; and CD45RA + CD27 -(terminally differentiated memory T cells (TEMRA)]. Activation was assessed by HLA-DR expression on CD4 + and CD8 + cells. Senescence on CD4 + and CD8 + cells was assessed by CD57 and CD28 expression (CD57 + CD28 -measuring senescent T lymphocytes).

Duffau P., Wittkop L., Lazaro E., le Marec F., Cognet C., Blanco P., Moreau J.F., Dauchy F.A., Cazanave C., Vandenhende M.A., Bonnet F., Thiebaut R, & Pellegrin I. (2015). Association of immune-activation and senescence markers with non-AIDS-defining comorbidities in HIV-suppressed patients. AIDS (London, England), 29(16).

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