Tissues were fixed in 10% neutral buffered formalin (Sigma) for 24 h and transferred to 70% ethanol. Tissues were embedded in paraffin, and 3–5 µm sections were processed for H&E staining, immunohistochemistry and co-immunofluorescence using standard protocols as previously described [30 (link)]. The following antibodies and kits were used: SPARC (R&D Systems, AF942, 1:100, RRID: AB_2286625 ), α-SMA (Dako, Clone 1A4, 1:250, RRID: AB_2335694), CC3 (Cell signaling, 9664L, 1:100, RRID: AB_2070042) and Ki67 (Thermo Scientific, RM-9106, 1:200, RRID: AB_2341197 ), CD31 (BD Biosciences, 553370, 1:100, RRID: AB_394816 ). Pictures were taken with 40× magnification with an Olympus DP27 camera and the Olympus cellSens Entry 1.12 software.
Clone 1a4
Manufactured by Cell Signaling Technology
Clone 1A4 is a mouse monoclonal antibody that recognizes the human CD3 epsilon chain. It is suitable for use in applications such as flow cytometry and immunohistochemistry.
Automatically generated - may contain errors
Lab products found in correlation
Cellsens entry 1, by Olympus (1 mentions)
Neutral buffered formalin, by Merck Group (1 mentions)
Dp27 camera, by Olympus (1 mentions)
Dako liquid dab substrate chromogen system, by Agilent Technologies (1 mentions)
Anti phospho mtor, by Cell Signaling Technology (1 mentions)
Anti α actin, by Abcam (1 mentions)
Anti il 10, by Abcam (1 mentions)
Anti ephb4, by Santa Cruz Biotechnology (1 mentions)
Af4117, by R&D Systems (1 mentions)
Anti mtor, by Cell Signaling Technology (1 mentions)
Anti cd34, by R&D Systems (1 mentions)
Anti cleaved caspase 3, by Cell Signaling Technology (1 mentions)
Anti ki67, by Abcam (1 mentions)
Anti gapdh, by Cell Signaling Technology (1 mentions)
Anti vegfr2, by Abcam (1 mentions)
Anti cd68, by Abcam (1 mentions)
Anti cd31, by Abcam (1 mentions)
Af942, by R&D Systems (1 mentions)
3 protocols using clone 1a4
1
Histological Analysis of Tissue Samples
2
Comprehensive Immunohistochemistry and Immunofluorescence Protocol
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Primary antibodies included: anti-α-actin (Abcam, ab5694; IHC and IF, 1:100; WB, 1:1000); Dako actin (Smooth Muscle) Clone 1A4; anti-cleaved Caspase-3 (Cell Signaling #9661; IHC, 1:50; WB, 1:1000); anti-CD31 (Abcam, ab28364; IHC and IF, 1:50); anti-CD34 (R&D, AF4117; IF, 1:100; WB, 1:1000); anti-CD68 (ED1; Abcam, ab31630; IHC, 1:200; WB, 1:1000); anti-Eph-B4 (Santa Cruz, sc-5536; IF, 1:50); anti-Ephrin-B2 (Novus, NBP1-48610; IHC, 1:50); anti-GAPDH (Cell Signaling, 14C10; WB, 1:2000); anti-IL-10 (Abcam, ab9969; IF, 1:100); anti-Ki67 (Abcam, ab15580; IHC and IF, 1:100; WB, 1:1000); anti-phospho-mTOR (Cell Signaling, #2971; IHC, 1,50; WB,1:1000); anti-mTOR (Cell Signaling, #2972; WB, 1:1000); anti-transglutaminase 2 (TGM2; #37557; IF, 1:100); anti-VEGFR2 (ABCAM, ab2349; IF, 1:100; WB,1:1000); Secondary antibodies used for IF were: donkey anti-goat Alexa-Fluor-488, donkey anti-rabbit Alexa-Fluor-488, donkey anti-rabbit Alexa-Fluor-568, donkey anti-mouse Alexa-Fluor-568 and chicken anti-mouse Alexa-Fluor-488 conjugated antibodies from Invitrogen (1:1000). For IHC, sections were incubated with EnVision reagents for 1 h at room temperature and treated with Dako Liquid DAB + Substrate Chromogen System (Dako). Finally, the sections were counterstained with Mayer’s hematoxylin.
3
Histological Analysis of Tissue Samples
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Tissues were fixed in 10% neutral buffered formalin for 24 h and transferred to 70% ethanol. Tissues were embedded in paraffin and 4 μm sections were processed for H&E staining, picrosirius staining, immunohistochemistry (IHC) and co‐immunofluorescence (Co‐IF) using standard protocol as previously stated.18 The following antibodies were used: SPARC (R&D Systems, AF942, 1:100, Research Resource Identifiers (RRID): Antibody (AB)_2286625), α‐SMA (Dako, Clone 1A4, 1:250, RRID: AB_2335694), CC3 (Cell signalling, 9664L, 1:100, RRID: AB_2335694) CCL2 (Invitrogen, #MA5‐17040, Clone 2D8, 1:500, RRID: AB_2538512), CD45 (BD Biosciences, 550539, 1:100, RRID: AB_2174426).
Pictures were taken with 40x magnification for H&E and IHC staining and 20x magnification for Co‐IF staining. Olympus DP27 and Olympus confocal FV1000 cameras were used for visualization.
Pictures were taken with 40x magnification for H&E and IHC staining and 20x magnification for Co‐IF staining. Olympus DP27 and Olympus confocal FV1000 cameras were used for visualization.
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