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Clone 4b11

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The Leica Clone 4B11 is a high-precision laboratory instrument designed for specific applications. It offers reliable and consistent performance in controlled environments.

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Lab products found in correlation

Clone ep459y, by Thermo Fisher Scientific (2 mentions) Anti rabbit and anti mouse hrp secondary antibodies, by Abcam (2 mentions) Dmi8 inverted microscope, by Leica (2 mentions) Clone ln10, by Leica (2 mentions) Clone fm264g, by BioLegend (2 mentions) Clone ps1, by Leica (2 mentions) Bond polymer refine detection kit, by Leica (1 mentions) Caseviewer software, by 3DHISTECH (1 mentions) Ventana benchmark ultra, by Roche (1 mentions) Tissue preparation system, by Siemens (1 mentions) Clone ep1, by Agilent Technologies (1 mentions) Pr clone pgr 636, by Agilent Technologies (1 mentions) Clone do 7, by Agilent Technologies (1 mentions)

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CD8 (clone 4B11, (16 citations) Anti-CD8 (clone 4B11; (4 citations)

10 protocols using clone 4b11

1

Quantifying GBM B Cell Infiltration

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A total of 60 GBM patients were analyzed for the presence of CD20+ B cells by IHC. Formalin-fixed paraffin-embedded tumor blocks of GBM patients (see Table 1 for patients’ characteristics) were double-labeled with rabbit anti-human CD20 (1:2 dilution, clone EP459Y, Thermo Fischer) and mouse anti-human CD8 (1:100 dilution, clone 4B11, Leica). Anti-rabbit and anti-mouse HRP secondary antibodies (both from Abcam) were used at 1:500 dilution. Twenty consecutive high-power fields were evaluated on each tumor by a board-certified neuropathologist from the Neurosurgical Department at Northwestern University. Images were captured on a Leica DMi8 inverted microscope.
Lee-Chang C., Rashidi A., Miska J., Zhang P., Pituch K.C., Hou D., Xiao T., Fischietti M., Kang S.J., Appin C.L., Horbinski C., Platanias L.C., Lopez-Rosas A., Han Y., Balyasnikova I.V, & Lesniak M.S. (2019). Myeloid-derived suppressive cells promote B cell–mediated immunosuppression via transfer of PD-L1 in glioblastoma. Cancer immunology research, 7(12), 1928-1943.
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2

Immunohistochemistry for CD8, CD4, and CD3

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Primary antibodies against human CD8 (Clone 4B11, Cat# PA0183), CD4 (Clone 4B12, Cat# PA0427) and CD3 (Clone LN10, Cat# PA0553) were purchased from Leica (Leica Biosystems, Newcastle Upon Tyne, UK), visualization of the antibody staining was performed with the recommended Bond™ Polymer Refine Detection kit (Leica Biosystems). PD-L1 IHC staining were performed with the 22C3 pharmDx kit (Cat# SK006; Dako Inc.). All these primary antibodies were prepared read-to-use according to the user's manuals.
Zhu L., Cheng G., Wu M., Chen M, & Jin Y. (2023). Heterogeneous distribution pattern of CD3+ tumor-infiltrated lymphocytes (TILs) and high combined positive score (CPS) favored the prognosis of resected early stage small-cell lung cancer. Translational Oncology, 34, 101697.
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3

Quantifying Tumor-Infiltrating Lymphocytes in GBM

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Formalin-fixed paraffin-embedded tumor blocks of 60 GBM patients were double-labeled with rabbit anti-human CD20 (1:2 dilution, clone EP459Y, Thermo Fischer) and mouse anti-human CD8 (1:100 dilution, clone 4B11, Leica). Anti-rabbit and anti-mouse HRP secondary antibodies (both from Abcam) were used at 1:500 dilution. Twenty consecutive high-power fields were evaluated on each tumor by a board-certified neuropathologist from the Neurosurgical Department at Northwestern University. Images were captured on a Leica DMi8 inverted microscope.
Hou D., Wan H., Katz J.L., Wang S., Castro B.A., Vazquez-Cervantes G.I., Arrieta V.A., Dhiantravan S., Najem H., Rashidi A., Chia T.Y., Arjmandi T., Collado J., Billingham L., Lopez-Rosas A., Han Y., Sonabend A.M., Heimberger A.B., Zhang P., Miska J, & Lee-Chang C. (2024). Antigen-presenting B cells promote TCF-1+ PD1- stem-like CD8+ T-cell proliferation in glioblastoma. Frontiers in Immunology, 14, 1295218.
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4

Immunohistochemical Analysis of CD8+ T Cells

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Tissue in paraffin blocks that was residual after histopathologic diagnoses had been finalized were used for immunohistochemical (IHC) staining for CD8. Five-micron sections were cut from formalin-fixed, paraffin-embedded tissue from the pre-vaccination (diagnostic biopsy) and from the post-vaccination resection (7 weeks after the third vaccination). Heat-based antigen retrieval was performed for 30 minutes, followed by blocking endogeneous peroxidase with 0.3% H2O2, and incubation with primary antibody. The primary antibody was detected using the PowerVision+ Poly-HRP IHC detection system (Leica Biosystems), as per the manufacturer’s instructions. After incubation with detection reagents, Diaminobenzidine was used as a chromogen, and Harris hematoxylin was used as a counterstain. The primary antibody used for CD8+ was clone 4B11 (Leica Biosystems, dilution 1:500, incubation overnight at 4°C).
Alvarez R.D., Huh W.K., Bae S., Lamb LS J.r., Conner M.G., Boyer J., Wang C., Hung C.F., Sauter E., Paradis M., Adams E.A., Hester S., Jackson B.E., Wu T.C, & Trimble C. (2015). A Pilot Study of pNGVL4a-CRT/E7(detox) for the Treatment of Patients with HPV16+ Cervical Intraepithelial Neoplasia 2/3 (CIN2/3). Gynecologic oncology, 140(2), 245-252.
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5

Evaluating CD8+ Cells in Neuroblastoma

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Previously constructed TMAs of neuroblastomas28 were evaluated for CD8 expression by immunohistochemistry. The TMA was constructed with duplicate 1.00 mm cores from each patient. Immunohistochemistry for CD8 was performed with an antibody purchased from Leica Biosystems (Clone 4B11, Buffalo Grove, IL). CD8 expression was scored as (0) none, (1) rare scatted CD8+ cells (less than 5), (2) occasional CD8+ cells (less than 10), (3) aggregates of CD8+ cells, 10 or more cells with only a single aggregate of 2 or more CD8+ cells within 10 μm proximity to each other, (4) multiple aggregates of CD8+ cells. The average of the two independent cores were taken and rounded up. Two-sided Wilcoxon Rank Sum test was used to compare MYCN-NA and MYCN-A tumors.
Wei J.S., Kuznetsov I.B., Zhang S., Song Y.K., Asgharzadeh S., Sindiri S., Wen X., Patidar R., Najaraj S., Walton A., Auvil J.M., Gerhard D.S., Yuksel A., Catchpoole D., Hewitt S.M., Sondel P.M., Seeger R., Maris J.M, & Khan J. (2018). Clinically Relevant Cytotoxic Immune Cell Signatures and Clonal Expansion of T Cell Receptors in High-risk MYCN-not-amplified Human Neuroblastoma. Clinical cancer research : an official journal of the American Association for Cancer Research, 24(22), 5673-5684.
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6

Tissue Culture of Resected Specimens

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As described previously75 (link), resected specimens were used for culture treatment. Briefly, the resected specimen rapidly was transferred into the lab in a sterile container, cut (size approximately 5 x 3 x 1 mm) and treated with respectively labelled and genetically modified cells. After culturing, the tissue was harvested and sectioned, subsequently being stained for human CD3epsilon (1:100 dilution, clone PS1, Novocastra, UK), CD8 (1:100 dilution, clone 4B11, Novocastra, UK) and for migrated cells a monoclonal anti-GFP antibody (clone FM264G, Biolegend) was used. Quantification was performed on whole slide sections as described previously76 (link). All material was obtained after approval by the medical ethics committee of the University of Heidelberg (S-069), written consent was obtained from all patients prior to analysis.
Lesch S., Blumenberg V., Stoiber S., Gottschlich A., Ogonek J., Cadilha B.L., Dantes Z., Rataj F., Dorman K., Lutz J., Karches C.H., Heise C., Kurzay M., Larimer B.M., Grassmann S., Rapp M., Nottebrock A., Kruger S., Tokarew N., Metzger P., Hoerth C., Benmebarek M.R., Dhoqina D., Grünmeier R., Seifert M., Oener A., Umut Ö., Joaquina S., Vimeux L., Tran T., Hank T., Baba T., Huynh D., Megens R.T., Janssen K.P., Jastroch M., Lamp D., Ruehland S., Di Pilato M., Pruessmann J.N., Thomas M., Marr C., Ormanns S., Reischer A., Hristov M., Tartour E., Donnadieu E., Rothenfusser S., Duewell P., König L.M., Schnurr M., Subklewe M., Liss A.S., Halama N., Reichert M., Mempel T.R., Endres S, & Kobold S. (2021). T cells armed with the C-X-C chemokine receptor type 6 enhance adoptive cell therapy for pancreatic tumours. Nature biomedical engineering, 5(11), 1246-1260.
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7

Quantifying CD8+ and CD3+ TILs in FFPE Tissue

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CD8 and CD3 IHC was performed for each case on a 4-mm tumor section cut from FFPE tissue blocks by using a monoclonal antibody against CD8 (clone 4B11,1 : 100 dilution, Novocastra, Newcastle, UK) and CD3 (clone LN10,1 : 100 dilution, Novocastra, Newcastle, UK) on an automated immunostainer. Case-viewer software (3DHISTECH.Ltd) was used to quantify total tumor area and percent of positive staining cells. The percentage of cells with weak, medium, and strong positive staining was determined using the color-deconvolution tool. CD8+ and CD3+ TILs density was calculated as the sum total of CD8+ and CD3+ cells (including those with weak, medium, and strong signals) divided by the area comprising the tumor and areas in direct contact with the tumor periphery (mm2).
Xu S.M., Shi C.J., Xia R.H., Wang L.Z., Tian Z., Ye W.M., Liu L., Liu S.L., Zhang C.Y., Hu Y.H., Zhou R., Han Y., Wang Y., Zhang Z.Y, & Li J. (2022). Analysis of Immunological Characteristics and Genomic Alterations in HPV-Positive Oropharyngeal Squamous Cell Carcinoma Based on PD-L1 Expression. Frontiers in Immunology, 12, 798424.
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8

Immunohistochemical Analysis of T-Cell Subsets

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Paraffin-embedded, 4-µm-thick skin tissue sections were stained for CD3 (RTU, clone 2GV6, Roche 790-4341), CD4 (1:100 clone SP35, Cell Marque, 104R-16), and CD8 (1:50, clone 4B11, Novocastra, NCL-L-CD8-4B11), using the Ventana Benchmark Ultra instrument (Roche). Tissue sections were pretreated in cell conditioning 1 buffer (pH 8.5) (Roche 950-124) at 98 °C for 64 min, followed by incubation with the antibodies at 37 °C for 44 min. The multimer-based detection kit, Ultraview (Roche 760-700) was used for the detection of CD3, and CD8. Optiview (Roche 760-700) was used to detect CD4. The reactions were visualized with diaminobenzidine (DAB) and counterstained with hematoxylin.
Fortino V., Wisgrill L., Werner P., Suomela S., Linder N., Jalonen E., Suomalainen A., Marwah V., Kero M., Pesonen M., Lundin J., Lauerma A., Aalto-Korte K., Greco D., Alenius H, & Fyhrquist N. (2020). Machine-learning–driven biomarker discovery for the discrimination between allergic and irritant contact dermatitis. Proceedings of the National Academy of Sciences of the United States of America, 117(52), 33474-33485.
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9

Tissue Culture of Resected Specimens

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As described previously75 (link), resected specimens were used for culture treatment. Briefly, the resected specimen rapidly was transferred into the lab in a sterile container, cut (size approximately 5 x 3 x 1 mm) and treated with respectively labelled and genetically modified cells. After culturing, the tissue was harvested and sectioned, subsequently being stained for human CD3epsilon (1:100 dilution, clone PS1, Novocastra, UK), CD8 (1:100 dilution, clone 4B11, Novocastra, UK) and for migrated cells a monoclonal anti-GFP antibody (clone FM264G, Biolegend) was used. Quantification was performed on whole slide sections as described previously76 (link). All material was obtained after approval by the medical ethics committee of the University of Heidelberg (S-069), written consent was obtained from all patients prior to analysis.
Lesch S., Blumenberg V., Stoiber S., Gottschlich A., Ogonek J., Cadilha B.L., Dantes Z., Rataj F., Dorman K., Lutz J., Karches C.H., Heise C., Kurzay M., Larimer B.M., Grassmann S., Rapp M., Nottebrock A., Kruger S., Tokarew N., Metzger P., Hoerth C., Benmebarek M.R., Dhoqina D., Grünmeier R., Seifert M., Oener A., Umut Ö., Joaquina S., Vimeux L., Tran T., Hank T., Baba T., Huynh D., Megens R.T., Janssen K.P., Jastroch M., Lamp D., Ruehland S., Di Pilato M., Pruessmann J.N., Thomas M., Marr C., Ormanns S., Reischer A., Hristov M., Tartour E., Donnadieu E., Rothenfusser S., Duewell P., König L.M., Schnurr M., Subklewe M., Liss A.S., Halama N., Reichert M., Mempel T.R., Endres S, & Kobold S. (2021). T cells armed with the C-X-C chemokine receptor type 6 enhance adoptive cell therapy for pancreatic tumours. Nature biomedical engineering, 5(11), 1246-1260.
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10

Molecular Analysis of Tumor DNA

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Cases were stained for p53 (clone DO-7, 1:2,000, DAKO), Wilms tumor 1 (WT-1, clone 6F-H1, 1:3,200, Invitrogen), estrogen receptor (ER, Clone EP1, 1:200, DAKO), progesterone receptor (PR, Clone Pgr636, 1:400, DAKO), and CD8 (Clone 4B11, 1:2,000, Novocastra). Procedures and scoring methods are described in the Supplementary Materials and Methods.
Molecular analysis DNA isolation. Tumor DNA was isolated from FFPE-tissue blocks either by using three 0.6-mm tumor cores (n ¼ 16) or by using microdissected tissue from 5 to 10 tissue sections (10 mm; n ¼ 26). DNA isolation was performed fully automated using the Tissue Preparation System (Siemens Healthcare Diagnostics) as described previously (28) . The median tumor cell percentage of the isolated areas was 80% (range, 25%-90%).
de Jonge M.M., Ritterhouse L.L., de Kroon C.D., Vreeswijk M.P.G., Segal J.P., Puranik R., Hollema H., Rookus M.A., van Asperen C.J., van Leeuwen F.E., Smit V.T.H.B.M., Howitt B.E, & Bosse T. (2019). Germline BRCA-Associated Endometrial Carcinoma Is a Distinct Clinicopathologic Entity. Clinical cancer research : an official journal of the American Association for Cancer Research, 25(24).
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