Immunostaining was previously performed on these TMAs using genetically validated rabbit monoclonal antibodies and staining protocols for ERG (EPR3864; Ventana) and PTEN (D4.3; Cell Signaling Technologies) on the Ventana Benchmark or Ventana Discovery Ultra (Ventana/Roche) (11 (link)). Visual scoring using a dichotomous and validated scoring system was performed (11 (link)).
Epr3864
Manufactured by Roche
Sourced in Australia, United States
The EPR3864 is a piece of laboratory equipment. It is a device designed for measuring and analyzing samples using electron paramagnetic resonance (EPR) spectroscopy. The core function of the EPR3864 is to detect and characterize the presence of unpaired electrons in materials, which can provide information about the chemical and physical properties of the sample.
Automatically generated - may contain errors
Lab products found in correlation
Ventana discovery ultra, by Roche (1 mentions)
Ventana benchmark, by Roche (1 mentions)
Benchmark platform, by Roche (1 mentions)
Anti cd31, by Agilent Technologies (1 mentions)
Pannoramic 1000 scanner, by 3DHISTECH (1 mentions)
Anti cd34, by Agilent Technologies (1 mentions)
Case center, by 3DHISTECH (1 mentions)
Anti pan cytokeratin, by Agilent Technologies (1 mentions)
Ventana discovery ultra system, by Roche (1 mentions)
Ab32072, by Roche (1 mentions)
Ab136650, by Abcam (1 mentions)
Pv6119, by Leica (1 mentions)
Ab32072, by Abcam (1 mentions)
Ventana ultra view universal dab detection kit, by Roche (1 mentions)
Ventana benchmark ultra system, by Roche (1 mentions)
6 protocols using epr3864
1
Immunostaining Validation for ERG and PTEN
2
Evaluating ERG Expression in Tumors
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
IHC was performed on sections that are selected for scanning for all in-house cases using anti-ERG rabbit monoclonal antibodies (EPR3864, Ventana, prediluted). Appropriate positive and negative controls were included. ERG immunohistochemical expression was performed based on clinically used evaluation criterion where expression of ERG protein within a tumor focus was considered to be positive and such a tumor focus was designated as ERG-positive. Tumor foci which do not expression ERG protein were designated as ERG-negative.
3
Bone-Implant Interface Vascularization Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The implant-bone samples were fixed in formalin for 2-7 days, decalcified in formic acid for 7 days, and embedded in paraffin. Three-micron sections were cut for immunohistochemistry and stained with CD34, collagen IV, and ETS-related gene (ERG) endothelial markers. ERG was chosen as the preferred antibody because collagen IV also stained the membranes of the fatty cells, making it difficult to interpret. ERG was easier to quantify because it only stains nuclei [32] , compared to CD34 and collagen IV, which stain the membrane and basal lamina [33, 34] .
ERG is a rabbit monoclonal antibody (EPR3864, Ventana Medical Systems) and runs on the Ventana Benchmark platform in a ready-to-use concentration.
NDP.view2 was used to view the scanned slides, and QuPath-0.2.3 was used for quantification of ERG-positive capillaries stained brown by the 3.3 ´-Diaminobenzidine (DAB) reaction.
Positive nuclei and the total number of nuclei were recorded in the 2-mm implant gap area (Figure 4). Unfortunately, the areas containing hydroxyapatite were difficult to section, resulting in lost tissue, which made comparison between groups impossible. Thus, immunohistochemical results were not included in this study.
(FIGURE 4 SHOULD BE PLACED HERE)
ERG is a rabbit monoclonal antibody (EPR3864, Ventana Medical Systems) and runs on the Ventana Benchmark platform in a ready-to-use concentration.
NDP.view2 was used to view the scanned slides, and QuPath-0.2.3 was used for quantification of ERG-positive capillaries stained brown by the 3.3 ´-Diaminobenzidine (DAB) reaction.
Positive nuclei and the total number of nuclei were recorded in the 2-mm implant gap area (Figure 4). Unfortunately, the areas containing hydroxyapatite were difficult to section, resulting in lost tissue, which made comparison between groups impossible. Thus, immunohistochemical results were not included in this study.
(FIGURE 4 SHOULD BE PLACED HERE)
4
Immunohistochemical Analysis of Tumor Samples
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
FFPE tumour samples were sectioned and stained with haematoxylin and eosin (H&E), or the following antibodies: anti-pan-cytokeratin (mouse, AE1/AE3, Dako, North Sydney, NSW, Australia), anti-CD31 (mouse, JC70A, Dako), anti-CD34 (mouse, QBEnd10, Dako), anti-ERG (rabbit, EPR3864, Roche Diagnostics, North Ryde, NSW, Australia), anti-CAMTA1 (rabbit, polyclonal, Novus Biologicals, Noble Park North, VIC, Australia), and anti-TFE3 (rabbit, EPR11591, Abcam, Melbourne, VIC, Australia). H&E and IHC slides were scanned digitally at 20× magnification using the Pannoramic 1000 scanner (3DHISTECH Ltd., Budapest, Hungary). High-definition images were uploaded into CaseCenter (3DHISTECH Ltd.), and images were processed using FIJI image analysis software 2.14.0 [46 (link)].
5
Chromogenic IHC on FFPE and Fresh Frozen Tissue
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
All single-plex DAB-based chromogenic IHC on tissue samples (FFPE or fresh frozen) was performed on a Ventana Discovery ULTRA system (Roche). Fresh frozen tissues were air dried at room temperature for 15 minutes, fixed overnight in formalin, and rinsed off with dH2O prior to staining. IHC labeling for c-Myc and ERG was performed with primary antibodies against c-MYC (1:600, Abcam, ab32072) and ERG (EPR3864) (Roche, 6478450001), respectively. IHC for EdU and MYC on cells grown on chamber slides was performed as described (48 (link)). A primary antibody against BrdU (Abcam, ab136650) was validated to be specific for EdU as well, and used at 1:1,000. The primary antibody against c-MYC was used at 1:250 dilution (Abcam, ab32072). The secondary antibodies were PowerVision Poly-HRP anti-rabbit or anti-mouse (Leica, PV6114 and PV6119, respectively) and developed with DAB.
6
Evaluating ERG Protein Expression in Prostate Cancer
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
From the TMAs, 480 patients had at least one malignant sample that could be evaluated for in situ ERG protein expression. In total, 2047 tissue cores (1447 malignant, 600 benign) could be evaluated, of which 1–11 malignant tissue cores (median: 3 tissue cores) and 0–6 benign tissue cores (median: 1 tissue core) from each patient. Multiple tissue cores from the same malignant tumor focus were available for 312 patients and from multiple foci from 156 patients.
In situ protein expression of ERG was assessed with immunohistochemistry on the TMAs using the fully automated Ventana Benchmark Ultra system, with anti‐ERG monoclonal antibody EPR3864 (Roche Tissue Diagnostics, Tucson, AZ, USA) and Ventana UltraView Universal DAB Detection kit (Roche Tissue Diagnostics). Tissue from tonsils, liver, pancreas and appendix were used as controls for ERG immunohistochemistry. Endothelial cells were used as an internal positive control. ERG protein expression was visually scored and evaluated as positive or negative as previously described [26 (link), 27 (link)]. A tissue core was classified as positive if any percentage of malignant cells showed positive nuclear staining.
In situ protein expression of ERG was assessed with immunohistochemistry on the TMAs using the fully automated Ventana Benchmark Ultra system, with anti‐ERG monoclonal antibody EPR3864 (Roche Tissue Diagnostics, Tucson, AZ, USA) and Ventana UltraView Universal DAB Detection kit (Roche Tissue Diagnostics). Tissue from tonsils, liver, pancreas and appendix were used as controls for ERG immunohistochemistry. Endothelial cells were used as an internal positive control. ERG protein expression was visually scored and evaluated as positive or negative as previously described [26 (link), 27 (link)]. A tissue core was classified as positive if any percentage of malignant cells showed positive nuclear staining.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!