UniProt Accession Number | Reagent Type | Target Name / Protein Biomarker | Target Species | Host Organism | Isotype | Clonality | Vendor | Catalog Number | Conjugate | RRID | Availability | Method | Tissue Preservation | Target Tissue | Tissue State | Detergent | Antigen Retrieval Conditions | Dye Inactivation Conditions | Recommend | Agree | Disagree | Contributor | Notes |
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
P20023 | Primary Antibody | CD21 | Human | Mouse | IgG1 | Bu32 | BioLegend | 354903 | PE | AB_2561406 | Stock | IBEX2D Manual | 1:4 Cytofix/Cytoperm Fixed Frozen | Lymph Node | NA | 0.3% Triton-X-100 | NA | 1 mg/ml LiBH4 15 minutes | Yes | 0000-0003-4379-8967 [2, 1] | NA | 0000-0003-4379-8967 | 1 |
P20023 | Primary Antibody | CD21 | Human | Rabbit | IgG | SP186 | Abcam | ab306325 | PE | NA | Stock | Cell DIVE-IBEX | FFPE | Lymph Node | Follicular Lymphoma | 0.3% Triton-X-100 | pH 6 for 30 minutes ER1 (AF9961) and pH 9 for 30 minutes ER2 (AF9640) using the Leica Bond | 1 mg/ml LiBH4 15 minutes | Yes | 0000-0003-4379-8967 | NA | 0000-0003-4379-8967 | 2 |
P20023 | Primary Antibody | CD21 | Human | Rabbit | IgG | SP186 | Abcam | ab306325 | PE | NA | Stock | Cell DIVE-IBEX | FFPE | Lymph Node | Follicular Lymphoma | 0.3% Triton-X-100 | pH 6 for 30 minutes ER1 (AF9961) using the Leica Bond | 1 mg/ml LiBH4 15 minutes | No | 0000-0003-4379-8967 | NA | 0000-0003-4379-8967 | 2 |
Publications
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A. J. Radtke et al., “IBEX: an iterative immunolabeling and chemical bleaching method for high-content imaging of diverse tissues”, Nat. Protoc., 17(2):378-401, 2022, doi: 10.1038/s41596-021-00644-9.
A. J. Radtke et al., “Accompanying dataset for: IBEX: An iterative immunolabeling and chemical bleaching method for high-content imaging of diverse tissues”, doi: 10.5281/zenodo.5244550.
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A. J. Radtke et al., “IBEX: A versatile multiplex optical imaging approach for deep phenotyping and spatial analysis of cells in complex tissues”, Proc Natl Acad Sci, 117(52):33455–33465, 2020, doi:10.1073/pnas.2018488117
Additional Notes
- Evaluated in human lymph nodes and shown to label follicular dendritic cells and B cells. Clone Bu32 has been shown to work in multiple conjugates for IBEX experiments (AF488 and AF532).
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Clone SP186 labels follicular dendritic cells in FFPE samples following antigen retrieval with pH 6 and amplification with a secondary antibody. This direct conjugate gives beautiful labeling after dual antigen retrieval (pH 6 and pH 9) but a speckled, less robust labeling pattern with pH 6 alone (shown in the two images below).
Antigen retrieval, pH 6 and pH 9 Antigen retrieval, pH 6