After perfusion with PBS to remove blood, lip tissue from tumor and control mice was dissected, fixed overnight in buffered 3.7% formalin at room temperature, transferred to sequential 70%, 80%, 90% and 100% methanol, and embedded in paraffin. Sections were stained with H&E and examined microscopically by 2 pathologists. Images were acquired using an Olympus BX53 microscope (Tokyo, Japan). Unstained serial sections were used for immunohistochemistry with the following antibodies: goat antibody against mouse cytokeratin 14 (sc‐17104, Santa Cruz Biotechnology, Texas, USA); rabbit antibody against mouse Ki67 (790‐4286, Roche, Basel, Switzerland); rabbit antibody against mouse γH2AX (#9718, Cell Signaling Technology, Danvers, USA); rabbit antibody against mouse p‐Akt (ab38449, Abcam, Cambridge, UK); rabbit antibody against mouse p‐Erk1/2 (#4370, Cell Signaling Technology); rabbit antibody against mouse p‐Erk1/2 (#9101, Cell Signaling Technology); rabbit antibody against mouse p‐4EBP1 (#2855, Cell Signaling Technology); rabbit antibody against mouse p‐S6 protein (#4858, Cell Signaling Technology); and rabbit antibody against mouse p‐GSK3 (#9331, Cell Signaling Technology). Immunohistochemistry was performed using the Ventana Discovery XT system (Roche Tissue Diagnostics K. K. Basel, Switzerland).
Ventana discovery xt system
Manufactured by Roche
Sourced in United States
The Ventana Discovery XT system is a fully automated slide-staining instrument designed for use in clinical and research laboratories. The system is capable of performing a variety of immunohistochemistry and in situ hybridization assays. It is capable of processing up to 30 slides simultaneously and can be customized to meet the specific needs of the laboratory.
Automatically generated - may contain errors
Lab products found in correlation
H2o2 reagent, by Roche (2 mentions)
Herceptest kit, by Agilent Technologies (2 mentions)
Hematoxylin 2, by Roche (2 mentions)
Bx53 microscope, by Olympus (1 mentions)
Rabbit antibody against mouse p erk1 2, by Cell Signaling Technology (1 mentions)
Chromamap dab detection kit, by Roche (1 mentions)
Pannoramic viewer software, by 3DHISTECH (1 mentions)
Optiview dab ihc detection steps, by Roche (1 mentions)
Vysis her2 cep17 fish probe kit, by Abbott (1 mentions)
M7240, by Agilent Technologies (1 mentions)
Pharmdx kit, by Agilent Technologies (1 mentions)
Ribo cc buffer, by Roche (1 mentions)
Bluing reagent, by Roche (1 mentions)
Protease 1, by Roche (1 mentions)
Fsx100, by Olympus (1 mentions)
Discovery dab map detection kit, by Roche (1 mentions)
11 protocols using ventana discovery xt system
1
Histological Analysis of Tumor Tissue
2
Immunohistochemical Analysis of Sternum Bone Fragments
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Bone fragments from sternum were harvested, fixed in 10% neutral buffered formalin, decalcified, and then paraffin embedded. Immunohistochemical staining on 2.5μM sections was carried out using the Ventana Discovery XT system (Roche). Sections were deparaffinized, hydrated, and loaded on the Discovery XT. Antigen retrieval was performed using citrate buffer solution. Endogenous peroxidases were quenched using H2O2 reagent (Ventana). Primary antibodies against myeloperoxidase and factor VIII (both Dako, rabbit polyclonal, diluted 1:300), and CD31 (Abcam; rabbit polyclonal, diluted 1:50), and CD274 (Cell Signaling; diluted 1:150, rabbit mAb recognizing endogenous levels of total PD-L1 protein), were incubated for 20min and detected using an anti-rabbit secondary antibody and the ChromaMap DAB detection kit (Ventana). Tissues were counterstained with hematoxylin and image analysis was performed using Pannoramic Viewer Software (3DHISTECH).
3
Immunohistochemical Staining of PDAC
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
IHC staining was carried out on 2.5-mmol/L sections of PDAC and normal tissues on a tissue microarray, using the Ventana Discovery XT system (Roche). Sections were deparaffinized, hydrated, and loaded on the Discovery XT. Antigen retrieval was performed using citrate buffer solution. Endogenous peroxidases were quenched using H 2 O 2 reagent (Ventana). The BAC2 mAb has previously been used to specifically
4
Immunohistochemical Analysis of FH and 2SC
5
Evaluating ICAM-1 and HER2 Expression in Tissues
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
ICAM-1 expression in human tissue was evaluated by IHC using an anti-ICAM-1 antibody (G-5, Santa Cruz Biotechnology). Scoring was performed by an independent pathologist blinded to the patients’ clinical information. Protein expression was interpreted by the weighted histoscore method (H score).80 (link) The intensity of protein expression was scored as 0 (negative), 1 (light brown), 2 (brown), or 3 (dark brown). The final score was calculated as follows: (0 × % of negative cells) + (1 × % of light brown cells) + (2 × % of brown cells) + (3 × % of dark brown cells). Tumors with an H score of equal to or more than 10 were defined as positive ICAM-1 expression, while tumors with a score of less than 10 were defined as negative ICAM-1 expression. HER2 expression was analyzed by IHC using a HercepTest kit (Dako, Denmark), fluorescence in situ hybridization (FISH) using a Vysis HER2/CEP17 FISH probe kit (Abbott, USA) and a Dako detection kit (Dako, Denmark), and silver-enhanced in situ hybridization (SISH) using the Ventana Discovery XT system (Ventana/Roche, USA) according to the manufacturers’ instructions. FISH and SISH scores were assessed by detecting the fluorescence signal in 50 malignant cell nuclei. HER2 positivity was defined as either HER2 IHC 3+ or IHC 2+ and FISH/SISH positive (HER2/CEP17 [centromere enumerator probe 17] ≥ 2).
6
Quantitative Immunohistochemical Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Expression levels of therapeutic targets such as HER2/neu, EGFR and MIB1 were determined using immunohistochemistry (IHC). The immunohistochemical stainings with anti-HER2/neu (A0785; 1:300, DAKO, Hamburg, Germany), anti-EGFR (pharmDx™-kit, DAKO) and anti-MIB1 (M7240; 1:100, DAKO) antibodies were performed on 3 μm consecutive sections using an automated slide processing system (Ventana DISCOVERY XT System, Ventana Medical Systems, Inc., Tucson, USA) in accordance with the manufacturer's instructions. IHC staining and analysis were performed as previously described [37 (link)]. MIB1 score was defined as Score 0 (0 - 20% of all nuclei were positive), Score 1 (20 – 50%), Score 2 (50 – 80 %) and Score 3 (80 - 100%).
7
Immunohistochemical Analysis of IGF-1 in Lungs and Livers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The lungs and livers in experiment 1 were immunostained to measure insulin growth factor 1 (IGF‐1) levels using the avidin‐biotin complex (ABC) method. All staining processes from deparaffinization to counterstaining with hematoxylin were performed automatically using the Ventana Discovery XT system (Ventana Medical Systems, Tucson, AZ, USA). Antigen retrieval was performed, using the ‘extended mode’ with RiboCC buffer (Ventana Medical Systems) and the IGF‐1 rabbit polyclonal antibody for mice (bs‐0014R; Bioss Antibodies, Woburn, MA, USA), were used at a 1:25 dilution ratio with a reaction time of 60 min.
8
HER2 Status Evaluation Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Surgically or endoscopically resected primary tumor tissues were formalin fixed and paraffin embedded prior to being analyzed by experienced pathologists at Yonsei University College of Medicine. We performed HER2 IHC analysis using the HercepTest TM Kit (Dako, Denmark), FISH analysis using the Vysis TM HER2/CEP17 FISH Probe Kit (Abbott, USA), and Dako Detection Kit (Dako Denmark), and SISH analysis using the Ventana Discovery XT system (Ventana/Roche, USA) according to the manufacturers' instructions. FISH and SISH scores were assessed by detecting the fluorescence signal in 50 malignant cell nuclei. HER2 positivity was defined as HER2 IHC 3? or IHC 2? and FISH/SISH positive [HER2/CEP17 (centromere enumerator probe 17) ratio C2] [5] .
9
HER2 Silver In Situ Hybridization Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
HER2 SISH was performed on the Ventana Discovery XT system (Ventana), according to the manufacturer's protocols. The HER2 DNA probe (Ventana) was denatured at 95 °C for 12 min and hybridized at 52 °C for 2 h. The CEP17 probe (Ventana) was denatured at 95 °C for 12 min and hybridized at 44 °C for 2 h. To visualize the HER2 probes, the slides were incubated with secondary antibodies conjugated with horseradish peroxidase (HRP), followed by HRP-catalyzed silver precipitation (black dots). For visualization of the CEP17 probes, a rabbit anti-DNP primary antibody was detected by a secondary antibody conjugated with alkaline phosphatase, and a naphthol/fast red stain was subsequently applied. The sections were then counterstained with hematoxylin II (modified Mayer hematoxylin) and a bluing reagent (Ventana).
Silver in situ hybridization (SISH) scoring was performed by counting the signals of 60 non-overlapping nuclei with clearly distinguishable borders, after which the mean gene copy number and HER2/CEP17 ratio were calculated by dividing the sum of the gene copy number by 60. HER2 amplification was defined as a HER2/CEP17 ratio greater than 2.0 [9, 12] .
Silver in situ hybridization (SISH) scoring was performed by counting the signals of 60 non-overlapping nuclei with clearly distinguishable borders, after which the mean gene copy number and HER2/CEP17 ratio were calculated by dividing the sum of the gene copy number by 60. HER2 amplification was defined as a HER2/CEP17 ratio greater than 2.0 [9, 12] .
10
Immunohistochemical Analysis of Breast Cancer Specimens
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Breast cancer specimens, including normal areas which were the margin tissue, were obtained from patients at the time of surgery. The archival specimens were obtained from the Zhongshan Hospital of Traditional Chinese Medicine Affiliated to Guangzhou University of Chinese Medicine. The sections (4-µm) were mounted onto silane-coated slides. Immunohistochemistry was automatically performed using the Ventana XT System Discovery® (Roche). Briefly, tissue sections were treated with protease I (Roche) at 37̊C for 16 min for antigen retrieval. The following primary antibody was used: KRT81 polyclonal antibody was diluted at 1:25 (Abcam), and sections were incubated at 37̊C for 32 min. The tissue sections were then incubated at 37̊C for 20 min with a universal secondary antibody (Affinity). The site of peroxidase binding was determined using the DISCOVERY DABMap Detection kit (Roche). Sections were then counterstained with Hematoxylin II (Roche) for microscopic examination. As a negative control, nonimmune γ-globulin was used instead of the antibody. The specimens were observed and photographed using a fluorescence microscope FSX100 (Olympus).
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!