Formalin-fixed paraffin-embedded blocks from 172 of the 456 patients in our cohort were sectioned and stained for immunohistochemistry from one of our cohorts. Anti-CD8 (clone 4B11; Leica Biosystems, Newcastle, UK) and anti-CD4 (clone 4B12; Leica Biosystems) antibodies were detected using the Bond Polymer Refine Detection System (Leica Biosystems) according to the manufacturer’s instructions. CD8+ T cell and CD4+ T cell counts were determined avoiding areas of necrosis. In cases of heterogeneity, CD8+ T cell and CD4+ T cell counts were estimated at the tumor front within the area of deepest invasion. A minimum of three random fields were examined to assess both the tumor center and the intraepithelial compartment. In cases of heterogeneity, the count that best represented the entire section was assigned, as previously described [22 (link)].
Anti cd8 clone 4b11
Manufactured by Leica
Sourced in United Kingdom
The Anti-CD8 (clone 4B11) is a monoclonal antibody that recognizes the CD8 antigen, which is expressed on the surface of T-lymphocytes. This antibody is commonly used in flow cytometry and immunohistochemistry applications to identify and quantify CD8-positive cells.
Automatically generated - may contain errors
Lab products found in correlation
Anti cd4 clone 4b12, by Leica (2 mentions)
Bond polymer refine detection system, by Leica (1 mentions)
Bond 3, by Leica (1 mentions)
Anti pd l1 clone e1l3n, by Cell Signaling Technology (1 mentions)
Bond polymer refine detection kit, by Leica (1 mentions)
Dako autostainer universal staining system, by Agilent Technologies (1 mentions)
Chemmate dako envision detection kit, by Agilent Technologies (1 mentions)
Anti cd3 clone ln10, by Leica (1 mentions)
Anti cd123 clone 7g3, by BD (1 mentions)
Anti cd163 clone 10d6, by Leica (1 mentions)
Bond rx automated stainer system, by Leica (1 mentions)
4 protocols using anti cd8 clone 4b11
1
Quantification of Tumor-Infiltrating T Cells
2
Immunohistochemical Analysis of Thyroid Cancer
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Four μm sections of formalin-fixed paraffin-embedded tissues of thyroid cancer were stained with Hematoxylin and Eosin stain, and underwent immunohistochemistry (IHC) analysis. IHC was carried out using the following primary antibodies: an anti PD-L1 (clone E1L3N; 1/200; Cell Signaling Technology, Inc., (CST), Danvers, MA, USA), anti BRAFV600E (clone VE1; 1/500; Spring Bioscience, Pleasanton, CA, USA), and anti CD8 (clone 4B11; ready to use; Leica BIOSYSTEMS Ltd., Newcastle, UK). We conducted IHC using a Bond Polymer Refine Detection kit (Leica Biosystems Ltd., Newcastle, UK) on an automated staining platform (BOND-III, Leica Biosystems Ltd., Newcastle, UK), following the manufacturer’s instructions [29 (link)].
3
Glioma Molecular Profiling and TILs Analysis
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We obtained a total of 1,149 glioma cases of the brain (619 GBM cases and 530 low-grade glioma [LGG] cases) with known mRNA expression data from TCGA database (https://gdc.cancer.gov/about-data/publications/pancanatlas and https://www.cbioportal.org/ ) [12 (link)]. Normal samples as well as tumor samples with missing data were excluded from analysis. The analysis was finally performed on 525 cases with both virtual histopathological slides and clinical data (from a total of 619 GBM samples). We present the raw data of our study in Supplementary Data 1.
Immunohistochemical staining was performed to evaluate the presence or absence of tumor-infiltrating lymphocytes (TILs) in GBM human tissue diagnosed at Hanyang University Guri Hospital. Haematoxylin and eosin (H and E)-stained slides were reviewed by at least two pathologists for each case (Min and Kim). In non-necrotic tissue with TILs, immunostaining for anti-CD3 (clone LN10 Leica Biosystems, Newcastle, UK), anti-CD8 (clone 4B11 Leica Biosystems, Newcastle, UK) and anti-CD4 (clone 4B12 Leica Biosystems was performed using the Dako Autostainer Universal Staining System (DakoCytomation, Carpinteria, CA, USA) and the ChemMate™ Dako EnVision™ Detection Kit.
Immunohistochemical staining was performed to evaluate the presence or absence of tumor-infiltrating lymphocytes (TILs) in GBM human tissue diagnosed at Hanyang University Guri Hospital. Haematoxylin and eosin (H and E)-stained slides were reviewed by at least two pathologists for each case (Min and Kim). In non-necrotic tissue with TILs, immunostaining for anti-CD3 (clone LN10 Leica Biosystems, Newcastle, UK), anti-CD8 (clone 4B11 Leica Biosystems, Newcastle, UK) and anti-CD4 (clone 4B12 Leica Biosystems was performed using the Dako Autostainer Universal Staining System (DakoCytomation, Carpinteria, CA, USA) and the ChemMate™ Dako EnVision™ Detection Kit.
4
Validating GE Profiles in cHL
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A bulk GE dataset (GSE132348) of 103 cHL patients from Gene-Expression Omnibus (GEO) was used to validate the results10 (link). It was obtained using the same panel and methods as for the discovery dataset, without the 30 extra custom genes. After two cycles of ABVD, patients were classified following interim FDG-PET (iPET) response, where positive is equivalent to U (4 or 5 on the Deauville-five-point scale), and negative is equivalent to F. Early response to ABVD assessed with iPET is prognostic for cHL patients and has been used to identify GE profiles related to outcome10 (link).
Selected markers were validated using IHC on TMAs constructed with duplicate 0.6-mm tissue cores from tumor-rich selected areas of archival FFPE tumor blocks. Primary antibodies were anti-PTPN2 (clone 2A1D1, Fisher Scientific), monoclonal anti-CD123 (clone 7G3, BD Pharmingen), anti-CD163 (clone 10D6, Leica Biosystems), anti-CD68 (clone 514H12, Leica Biosystems), anti-CD8 (clone 4B11, Leica Biosystems), and anti-granzyme B (GrB) (clone 11F1, Leica Biosystems). Staining was done with the BOND RX automated Stainer system (Leica Biosystems). IHC counts were manually scored by one of the authors (J.L.S.R.). Finally, the most significant variables from ssGSEA that were correlated with a bad prognosis were included in a prognostic validation model.
Selected markers were validated using IHC on TMAs constructed with duplicate 0.6-mm tissue cores from tumor-rich selected areas of archival FFPE tumor blocks. Primary antibodies were anti-PTPN2 (clone 2A1D1, Fisher Scientific), monoclonal anti-CD123 (clone 7G3, BD Pharmingen), anti-CD163 (clone 10D6, Leica Biosystems), anti-CD68 (clone 514H12, Leica Biosystems), anti-CD8 (clone 4B11, Leica Biosystems), and anti-granzyme B (GrB) (clone 11F1, Leica Biosystems). Staining was done with the BOND RX automated Stainer system (Leica Biosystems). IHC counts were manually scored by one of the authors (J.L.S.R.). Finally, the most significant variables from ssGSEA that were correlated with a bad prognosis were included in a prognostic validation model.
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