Briefly, 4 μm tissue slices were prepared from cryopreserved tissues and mounted on glass slides. Immunohistochemistry was performed according to the manufacturer’s instructions using an automated staining facility (Bond Max, Leica, Wetzlar, Germany). The following mouse monoclonal antibodies were utilized: CD3ϵ (clone ab16669, 1:100, Abcam, Cambridge, UK; RRID: AB_443425), CD4 (clone 4B12, 1:150, Leica, Wetzlar, Germany; RRID: AB_563559), CD8 (clone 4B11, 1:100, Leica, Wetzlar, Germany; RRID: AB_442068), CD20 (clone L26, 1:100, Leica, Wetzlar, Germany; RRID: AB_442055), CD163 (clone EPR19518, 1:500, Abcam, Cambridge, UK; RRID: AB_2753196), NKp46 (clone 195314, 1:175, R&D Systems, Minneapolis, MN, USA; RRID: AB_2149153) and FoxP3 (clone 236A/E7, 1:100, Thermo Fisher Scientific, Waltham, MA, USA; RRID: AB_467555).
Cd20 clone l26
Manufactured by Leica
Sourced in Germany
The CD20 (clone L26) is a laboratory equipment product used for the identification and quantification of B lymphocytes in various biological samples. It serves as a cell surface marker for B cells, providing a tool for research and diagnostic applications.
Automatically generated - may contain errors
Lab products found in correlation
Cd8 clone 4b11, by Leica (3 mentions)
Nkp46 clone 195314, by R&D Systems (3 mentions)
Cd4 clone 4b12, by Leica (3 mentions)
Ki 67 clone mib 1, by Agilent Technologies (3 mentions)
Cd3 clone ab16669, by Abcam (2 mentions)
Cd163 clone epr19518, by Abcam (2 mentions)
Foxp3 clone 236a e7, by Thermo Fisher Scientific (2 mentions)
Bond max, by Leica (2 mentions)
Pd l1 clone e1l3n, by Cell Signaling Technology (1 mentions)
Anti cd3 clone ln10, by Leica (1 mentions)
Mlh1 clone es05, by Leica (1 mentions)
Pms2 clone m0r4g, by Leica (1 mentions)
Ventana benchmark, by Roche (1 mentions)
Cd68 clone kp1, by Agilent Technologies (1 mentions)
Foxp3 clone 236a e7, by Abcam (1 mentions)
Msh2 clone fe11, by Agilent Technologies (1 mentions)
Y5200, by Agilent Technologies (1 mentions)
Ndp view2, by Hamamatsu Photonics (1 mentions)
Cd10 clone 56c6, by Leica (1 mentions)
Clone gi191e a8, by Cell Marque (1 mentions)
6 protocols using cd20 clone l26
1
Automated Immunohistochemistry Staining Protocol
2
Immunoprofiling of Pancreatic Ductal Adenocarcinoma
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The following mouse monoclonal antibodies were used to stain serial sections (4 µm) of cryopreserved PDA tissues and/or FFPE 3D bioprints: CD3ϵ (clone ab16669, 1:100, Abcam, UK; RRID: AB_443425), CD4 (clone 4B12, 1:150, Leica, Germany; RRID: AB_563559), CD8 (clone 4B11, 1:100, Leica, Germany; RRID: AB_442068), CD20 (clone L26, 1:100,Leica, Germany; RRID: AB_442055), CD163 (clone EPR19518, 1:500, Abcam, UK; RRID: AB_2753196), NKp46 (clone 195314, 1:175, R&D Systems, USA; RRID: AB_2149153), FoxP3 (clone 236A/E7, 1:100, Thermo Fisher Scientific, Germany; RRID: AB_467555), IL9 (clone EPR23484-151, 1:100, Abcam, UK), IL18 (clone EPR19954-188, 1:100, Abcam, UK), Granzyme B (clone 23H8L20, 1:200, Thermo Fisher Scientific, USA), Ki67 (clone MIB-1, 1:200, Dako, USA), LCK (clone 3A5, 1:50, Santa Cruz Biotechnology, USA). The complete staining procedure was carried out on a fully automated staining system (Bond-Max, Leica, Germany).
3
Comprehensive Immunophenotyping of FFPE Samples
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FFPE blocks were obtained from the Pathology Department of Tokyo Metropolitan Bokutoh Hospital. Immunohistochemistry was performed using the Ventana BenchMark automated immunostainer (Ventana Medical Systems, Tucson, AZ, USA) with labeled streptavidin–biotin and visualized with 3,3′-diaminobenzidine. The primary antibodies used were anti-CD3 (clone LN10, Leica), -CD4 (clone SP35, Ventana), -CD8 (clone 4B11, Leica), -CD45RO (clone UCHL-1, Ventana), -FOXP3 (clone 236A/E7, Abcam), -CD20 (clone L26, Leica), -NKp46 (clone #195314, R&D), -CD68 (clone Kp-1, Dako), -CD163 (clone 10D6, Leica), -CD204 (clone SRA-E5, Transgenic), -Ki-67 (clone MIB-1, Dako), -PD-L1 (clone E1L3N, Cell Signaling), -MLH1 (clone ES05, Leica), -MSH2 (clone FE11, Dako), -MSH6 (clone Polyclonal (Rabbit), GeneTex) and -PMS2 (clone M0R4G, Leica). EBV-encoded small RNA in situ hybridization (EBER-ISH) was performed on paraffin sections using a fluorescein isothiocyanate (FITC)-labeled peptide nucleic acid probe (Y5200; Dako, Glostrup, Denmark) and anti-FITC antibody (V0403, Dako). Slides were digitized with a Nanozoomer 2.0-HT virtual slide scanner (Hamamatsu Photonics, Hamamatsu, Japan) and observed in the NDP.view2 software (Hamamatsu Photonics). The density of immune cells was analyzed by Tissue Studio 2.0 software (Definiens, Munich, Germany).
4
Immunohistochemical Profiling of Follicular Lymphoma
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All cases were reviewed, and based on the revised 4th edition of the World Health Organization classification criteria, patients with testicular FL, duodenal-type FL, and pediatric-type FL were excluded [2 ]. The following pathological parameters were recorded: histologic grade (grades 1–2, grade 3A, and grade 3B), histologic pattern (follicular, follicular and diffuse, and diffuse), and immunophenotype (expression of BCL2, BCL6, CD10, or Ki-67). Immunohistochemical (IHC) staining was performed on whole slides using a fully automated IHC assay on a Ventana BenchMark XT Autostainer (Ventana Medical Systems, Woonsocket, RI, USA). Antibodies specific for the following markers were used: CD3 (clone PS1, mouse mAb, Novocastra, Newcastleupon-Tyne, UK), CD5 (clone 4C7, mouse mAb, Novocastra), CD20 (clone L26, mouse mAb, Novocastra), Ki-67 (mouse mAb, Dako, Glostrup, Denmark), CD10 (clone 56C6, mouse mAb, Novocastra), CD21 (clone 2G9, mouse mAb, Novocastra), BCL2 (clone E17, rabbit mAb, Cell Marque, Rocklin, CA, USA), and BCL6 (clone GI191E/A8, mouse mAb, Cell Marque). Immunostained slides from 239 FL cases were reviewed, and immunopositivity was defined as protein expression by ≥30% of the tumor cells [15 (link)-18 (link)]. Ki-67 proliferation index (PI) was assessed manually in neoplastic follicles, and a Ki-67 PI ≥ 30% was deemed as high expression [19 (link)-23 (link)].
5
STING Expression Analysis in Tumor Microenvironment
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Sections from tissue blocks of all cases were immunohistochemically stained with STING (anti-TMEM173; clone SP338, dilution 1:150; Abcam, UK). Heat-induced antigen retrieval for STING was performed using a microwave oven and 0.01mol/L of citrate buffer, pH 8.0, for 30 min. Moreover, tumors defined as “excluded” and “inflamed” by hematoxylin and eosin were investigated for the specific composition of different lymphocytes subpopulations, according to the different percentages of expression of the following antibodies: CD3 (clone PS1, dilution 1:200, LEICA), CD20 (clone L26, prediluted, NOVOCASTRA), CD4 (clone 4B12, dilution 1.150, LEICA), CD8 (clone 29S, dilution 1:20, LEICA), and FOXP3 (clone 221D/D3, dilution 1:200, SEROTEC). All samples were processed using a sensitive “Bond Polymer Refine” detection system in an automated bond immunohistochemistry instrument (Leica Biosystems, Germany). Sections incubated without the primary antibody served as a negative control. Cytoplasmic and membranous labeling for the STING was recorded by combining the percentage of positive cells (0–100%) multiplied by staining intensity (0, 1+, 2+, and 3+) to obtain an overall H-score (0-300).
6
Immunohistochemical Evaluation of Lymphoid Follicles
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To accurately evaluate the structures of the lymphoid follicles, including the germinal center and marginal zone, CD3 (polyclonal, Dako, CA, USA), CD20 (clone L26, Novocastra, Leica Biosystems, Germany), and Ki-67 (clone MIB1, Dako, CA, USA) immunohistochemical staining was performed on whole sections of formalin-fixed paraffin-embedded blocks in all cases. Details of immunohistochemical staining are provided in Supplementary Table 1 .
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