FACS analysis of T-cell subsets was performed according to a routine protocol. In brief, erythrocytes were lysed and cells were incubated with antibodies against CD3 (Becton Dickinson, San Diego, USA), CD4, CD8, CD28, CD45 (all Beckman Coulter, Brea; USA), CD45RA (Becton Dickinson) and CD45RO (Beckmann Coulter). For intracellular staining of Ki-67 (Becton Dickinson), cells were permeabilised with IntraPrep Reagents (Beckman Coulter) and measured within 2 h. All analyses were performed using Cytomics FC500 (Beckmann Coulter) and using CxP software.
Ki 67
Manufactured by BD
Sourced in United States
Ki-67 is a protein that is expressed during all active phases of the cell cycle, making it a useful marker for cell proliferation. It is commonly used in immunohistochemistry and flow cytometry applications to assess the growth fraction of a given cell population.
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2 protocols using ki 67
1
Flow Cytometry Analysis of T-Cell Subsets
2
FACS Analysis of T and B-cell Subsets
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FACS analysis of T and B-cell subsets was performed as previously described [15 (link)]. In brief, erythrocytes were lysed and leukocytes were incubated with antibodies against CD3 (Becton Dickinson, San Diego, USA), CD4, CD8, CD28, CD31, CD45, CD45RO (all Beckman Coulter, Brea; USA), CD19, CD197/CCR7, CD127, CD183/CXCR3, CD196/CCR6, CD194/CCR4 (all Miltenyi biotech, Bergisch Gladbach, Germany) and CD38 and CD45RA (Becton Dickinson) for T-cell phenotyping and CD19, CD21, CD24, CD27, CD38 IgD and IgM (all Miltenyi biotech) for B-cell phenotyping. For intracellular staining of Ki-67 (Becton Dickinson), cells were permeabilized with IntraPrep Reagents (Beckman Coulter; Brea, USA) and measured within 2 h. These analyses were performed on a FACS Canto II (Becton Dickinson) using FACS Diva and FlowJo software (Becton Dickinson).
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