After digesting, BMSCs were resuspended in PBS (106 cells/ml). Then, cell suspension (500 μl) was incubated with anti-CD44 antibody (1:40, Abcam) or anti-CD45 antibody (1:20, Abcam) in the dark for 30 min at room temperature. Subsequently, cells were resuspended in PBS (500 μl). The cellular phenotype of BMSCs was detected using flow cytometry.
Anti cd45 antibody
Manufactured by Abcam
Sourced in United States, United Kingdom
The Anti-CD45 antibody is a tool used in immunology research. It specifically binds to the CD45 protein, which is expressed on the surface of most hematopoietic cells. This antibody can be used to identify and isolate cells that express CD45, such as leukocytes.
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Lab products found in correlation
4 protocols using anti cd45 antibody
1
Immunophenotyping of Bone Marrow MSCs
2
Measuring Calcium Signaling in Leukocyte Subsets
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Isolated neutrophils, or total leukocytes from human buffy-coats were pelleted and resuspended in cell loading medium (KRG supplemented with Ca2+ with 1% heat-inactivated FCS) at 1 × 107 cells/ml. Cells were loaded with Fluo-3 AM (3.6 µl/ml) and Fura-Red (10 µl/ml) and incubated for 30 min at 37°C, washed twice and resuspended in cell-loading medium at 6 × 106 cells/ml. Before analysis, cells were incubated for 5 min at 37°C (with/without GPCR receptor antagonist (Table 1
When the total leukocyte population was used, cells were, after labeling with calcium dyes, also stained with anti-CD45 antibody (Abcam, diluted 1:500) for 20 min, at 4°C. CD45 is differentially expressed on distinct leukocyte subsets and facilitate the gating on neutrophils, monocytes and lymphocytes. The cell types (neutrophils, monocytes and lymphocytes) were gated based on CD45 and side scatter properties and the Ca2+ responses of the different cell types were analyzed separately (Figure 6B
When the total leukocyte population was used, cells were, after labeling with calcium dyes, also stained with anti-CD45 antibody (Abcam, diluted 1:500) for 20 min, at 4°C. CD45 is differentially expressed on distinct leukocyte subsets and facilitate the gating on neutrophils, monocytes and lymphocytes. The cell types (neutrophils, monocytes and lymphocytes) were gated based on CD45 and side scatter properties and the Ca2+ responses of the different cell types were analyzed separately (
3
Immunohistochemical Analysis of CD45 in OC
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Immunohistochemistry (IHC) staining was employed for the detection of CD45 expression in OC tissues. In brief, paraffin-embedded tissues were cut into 4-μm sections and subjected to deparaffinization with xylene and rehydration in a graded alcohol series, followed by incubation in an oven (65 °C) for 40 min (min). Sequential endogenous peroxidase blocking and antigen retrieval were carried out. Tissue slices were incubated with anti-CD45 antibody (1:100, Abcam, USA) in a humid chamber at 4 °C for 12 h (h). Subsequently, the slices were treated with anti-rabbit secondary antibody and incubated at 37 °C for 30 min, incubated with streptavidin-horseradish peroxidase conjugate, and developed with DAB. Then, the slices were stained with Mayer's hematoxylin, dehydrated, sealed with neutral size, and visualized under an Olympus CX31 microscope (Olympus, Center Valley, PA). Two pathologists blinded to the clinical characteristics of the OC patients independently evaluated the IHC results using the H-score system. CD45-positive cell intensity was assigned with a score of 0 (negative), 1 (weak), 2 (moderate), or 3 (strong). The percentage (0–100%) of stained cells was multiplied by the dominant intensity pattern of staining (0–3), and the H-score values ranged from 0 to 300. The expression of CD45 was dichotomized based on the median value.
4
Immune Cell Profiling by Flow Cytometry
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The rate of CD4+CD25+ T cells, NK cells, and T cells subgroup were all measured by flow
cytometry. The following antibodies were used in our study: anti-CD4 antibody (Abcam,
Cambridge, UK), anti-CD25 antibody (Abcam, Cambridge, UK), anti-CD127 antibody (Abcam,
Cambridge, UK), anti-CD3 antibody (Abcam, Cambridge, UK), anti-CD16 antibody (Abcam,
Cambridge, UK), anti-CD56 antibody (Abcam, Cambridge, UK), anti-CD8 antibody (Abcam,
Cambridge, UK), and anti-CD45 antibody (Abcam, Cambridge, UK). Flow cytometry analysis was
performed on a FACScan (BD Biosciences, Mountain View, California) using CellQuest
software.
cytometry. The following antibodies were used in our study: anti-CD4 antibody (Abcam,
Cambridge, UK), anti-CD25 antibody (Abcam, Cambridge, UK), anti-CD127 antibody (Abcam,
Cambridge, UK), anti-CD3 antibody (Abcam, Cambridge, UK), anti-CD16 antibody (Abcam,
Cambridge, UK), anti-CD56 antibody (Abcam, Cambridge, UK), anti-CD8 antibody (Abcam,
Cambridge, UK), and anti-CD45 antibody (Abcam, Cambridge, UK). Flow cytometry analysis was
performed on a FACScan (BD Biosciences, Mountain View, California) using CellQuest
software.
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