Routine blood counts and immunological phenotypes were evaluated as previously reported (Wang et al., 2018 (link)). The levels of serum immunoglobulins, including IgG, IgA, IgM, and IgE, were detected using nephelometry, while lymphocyte subsets were analyzed using a FACSCalibur flow cytometer (Becton Dickinson, Franklin Lakes, NJ, United States). The following validated antibodies were used for flow cytometry: anti-CD3 (UCHT1), anti-CD4 (RPA-T4), anti-CD8 (RPA-T8), anti-CD27 (M-T271), anti-TCRαβ (T10B9.1A-31), anti-TCRγδ (B1), anti-CD45RA (HI100), anti-CD19 (HIB19), anti-CD24 (ML5), anti-CD38 (HIT2), and anti-IgD (IA6–2) (all from BD Biosciences, San Jose, CA, United States).
Anti igd ia6 2
Manufactured by BD
Sourced in United States
Anti-IgD (IA6–2) is a monoclonal antibody that recognizes the IgD immunoglobulin. It is designed for use in laboratory research applications.
Automatically generated - may contain errors
Lab products found in correlation
Flow cytometry, by BD (2 mentions)
Facscalibur flow cytometer, by BD (1 mentions)
Anti cd8, by BD (1 mentions)
Lymphoprep, by STEMCELL (1 mentions)
Anti cd14 61d3, by Thermo Fisher Scientific (1 mentions)
Canto 2, by BD (1 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (1 mentions)
Facscalibur ow cytometer, by BD (1 mentions)
Facsdiva software, by BD (1 mentions)
5 protocols using anti igd ia6 2
1
Immunological Profiling of Blood Samples
2
Immune Cell Profiling Protocol
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Routine blood counts and immunological function analyses were performed. We used nephelometry to detect immunoglobulins, including IgG, IgA, and IgM, and lymphocyte subsets were measured using flow cytometry (Becton Dickinson, Franklin Lakes, NJ, USA). The following validated antibodies were used for flow cytometry: anti-CD3 (UCHT1), anti-CD8 (RPAT8), anti-CD27 (M-T271), anti-CD45RA (HI100), anti-CD4 (RPA-T4), anti-TCRαβ (T10B9.1A-31), anti-TCRγδ (B1), anti-CD19 (HIB19), anti-CD24 (ML5), anti-CD38 (HIT2), and anti-IgD (IA6–2) (all from BD Biosciences).
3
Isolation and Characterization of PBMCs
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Heparinized blood was collected from the patient and control subjects to isolate peripheral blood mononuclear cells using Lymphoprep (Stemcell Technologies). Control subjects were included using the following criteria: women of Caucasian descent between 20 and 35 years with no known (auto‐)immune diseases, recent infections (<30 days), or pregnancy. This study has been approved by the ethics committee (UZ Leuven, ref. nr. S60344) and has therefore been performed in accordance with the ethical standards laid down in the 1964 Declaration of Helsinki and its later amendments. All involved persons gave their informed consent prior to their inclusion in the study. PBMCs were frozen in 10% dimethyl sulfoxide (Sigma) in combination with fetal bovine serum and stored in liquid nitrogen. Thawed PBMCs were stained in phosphate‐buffered saline (PBS) supplemented with 3% fetal bovine serum (FBS) for live‐dead and surface markers for 1 h at 4°C. Anti‐human antibodies included are anti‐CD3 (REA613) (Miltenyi Biotec); anti‐CD14 (M5E5), anti‐CD27 (M‐T271) (both from BioLegend); anti‐IgD (IA6–2) (BD Biosciences); anti‐CCR7 (3D12), anti‐CD45RA (HI 100), anti‐CD8 (SK1), anti‐CD4 (RPA‐T4), anti‐CD3 (UCHT1), anti‐CD19 (HIB19) and anti‐CD14 (61D3) (all from eBioscience). Data were collected on BD CantoII (BD Biosciences) and analysed using FlowJo for Mac version 10.7 (BD Biosciences).
4
Comprehensive Immunological Profiling
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Routine blood counts and immunological function analyses were performed. We used nephelometry to detect immunoglobulins (Igs), including IgG, IgA, and IgM. Lymphocyte subsets were measured using flow cytometry (Becton Dickinson, Franklin Lakes, NJ, USA). The following validated antibodies were used for flow cytometry: anti-CD3 (UCHT1), anti-CD8 (RPAT8), anti-CD27 (M-T271), anti-CD45RA (HI100), anti-CD4 (RPA-T4), anti-TCRαβ (T10B9.1A-31), anti-TCRγδ (B1), anti-CD19 (HIB19), anti-CD24 (ML5), anti-CD38 (HIT2), and anti-IgD (IA6-2) (all from BD Biosciences) (Ding et al., 2018 (link)). flow cytometry detection of CD119 and CD212 was performed.
5
Immunoglobulin and Lymphocyte Subset Analysis
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The immunoglobulins, including IgG, IgA, and IgM, were detected by nephelometry, while the analyses of lymphocyte subsets were performed on a FACSCalibur ow cytometer (Becton Dickinson, Franklin Lakes, NJ, USA) and FACSDiva software (BD Biosciences). Brie y, samples were processed using the standard 'stain -lyse -wash' technique. The peripheral blood mononuclear cells (PBMCs) were stained for surface antigens with a panel of monoclonal antibodies and isotype control antibodies followed by red blood cell lysis (BD FACS) and washing with PBS. The following validated antibodies were used for ow cytometry: anti-CD3 (UCHT1), anti-CD8 (RPAT8), anti-CD27 (M-T271), anti-CD45RA (HI100), anti-CD4 (RPA-T4), anti-TCRαβ (T10B9.1A-31), anti-TCRγδ (B1), anti-CD19 (HIB19), anti-CD24 (ML5), anti-CD38 (HIT2), and anti-IgD (IA6-2) (all from BD Biosciences).
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