M7165

Manufactured by Agilent Technologies

Sourced in United States, Denmark

The M7165 is a high-performance microwave signal generator produced by Agilent Technologies. It is designed to generate precise and stable microwave signals across a wide frequency range. The M7165 offers a variety of features to support various applications in research, development, and testing environments.

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Lab products found in correlation

Ab37305, by Abcam (2 mentions) Streptavidin peroxidase, by Vector Laboratories (2 mentions) Human cd44h 2c5, by R&D Systems (2 mentions) Mouse igg and rabbit igg, by Agilent Technologies (1 mentions) Lsab2 system, by Agilent Technologies (1 mentions) M0851, by Agilent Technologies (1 mentions) Cx21 microscope, by Olympus (1 mentions) Ram34, by Thermo Fisher Scientific (1 mentions) Ab131259, by Abcam (1 mentions) Goat anti mouse igg, by Thermo Fisher Scientific (1 mentions) A11001, by Thermo Fisher Scientific (1 mentions) Goat anti rabbit igg, by Thermo Fisher Scientific (1 mentions) Evos fluorescence imaging system, by Thermo Fisher Scientific (1 mentions) Aperio cs2, by Leica (1 mentions) Ventana discovery xt, by Roche (1 mentions) M0823, by Agilent Technologies (1 mentions) M7018, by Agilent Technologies (1 mentions) M0888, by Agilent Technologies (1 mentions) Dako autostainer, by Agilent Technologies (1 mentions) Automated stainer, by Roche (1 mentions)

14 protocols using m7165

Quantitative Assessment of Angiogenesis

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The intensity of angiogenesis was measured quantitatively using MVD calculation. MVD was assessed on CD34-stained slides (M7165, DAKOCytomation, Denmark, 1:50, pH 9) by light microscopy in the areas having the highest number of capillaries with the lumen < 10 µm and small venules without lumen or with the lumen of < 10 µm (“hot spots”). Subsequently, only the microvessels were counted in three chosen fields in each sample (number of vessels/mm²) with a magnification of × 200. The MVD was calculated as the mean vessel count from these three fields, as described previously (Vermeulen et al. 1996 (link)). The CD34-positive blasts were considered as cells with nucleoli, on contrary to microvessels with lumen (hematoxylin staining) (Fig. 1).
Fig. 1

The estimation of microvessel density (MVD) using immunostaining of anti-CD34 antibody: a the areas having the highest number of capillaries and small venules (“hot spots”) were selected in low-power field (× 40) b then the microvessels were counted in these fields with a magnification of × 200: AML case (a, b) and control bone marrow trephine biopsy slides (c, d). The CD34-positive blast was considered as cells with nucleoli, on contrary to microvessels with lumen (hematoxylin staining). Magnification × 200

Gołos A., Jesionek-Kupnicka D., Gil L., Braun M., Komarnicki M., Robak T, & Wierzbowska A. (2019). The Expression of the SLIT–ROBO Family in Adult Patients with Acute Myeloid Leukemia. Archivum Immunologiae et Therapiae Experimentalis, 67(2), 109-123.

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Quantitative Analysis of Tumor Angiogenesis

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Paraffin sections were stained with monoclonal anti-CD34 antibody (1 : 100, M7165; Dako) and polyclonal anti-VEGF antibody (1 : 100, A20; Santa Cruz) by the streptavidin-biotin-peroxidase method (Dako LSAB2 System) as described previously [16 (link)]. The same concentrations of chromatographically purified mouse IgG and rabbit IgG (Dako) were used as negative controls. CD34-stained sections were evaluated for MVD, and vascular architecture parameters including vessel area, diameter, perimeter, and roundness were analyzed by using an image analyzer system (WinROOF, Mitani Corporation, Japan); WinROOF is an integrated software system for image processing, measurement, and data processing to support all image analysis-related operations [17 (link), 18 (link)]. The defined area for the measurement of these parameters was 1.0 mm². CD34-stained fields of 1.0 mm² (Figure 1(a)) were input into the image analyzer, and each vessel contour was manually traced in order to measure vessel density (number), vessel area (%), vessel diameter (μm), vessel perimeter (μm), and vessel roundness (0-1; 1 = completely round) (Figure 1(b)). Three different fields in each case were measured, and the median values were used for analysis. VEGF immunohistochemistry was defined as positive if more than 10% of adenoma cells were positive.

Takano S., Akutsu H., Hara T., Yamamoto T, & Matsumura A. (2014). Correlations of Vascular Architecture and Angiogenesis with Pituitary Adenoma Histotype. International Journal of Endocrinology, 2014, 989574.

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Immunohistochemical Analysis of α-SMA and CD34

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Multiple specimens were fixed in 10% buffered formalin, embedded in paraffin, and cut to 4 μm thickness. Our previous study presented the well-established routine protocol for immunohistochemical work in detail [12 (link)].
The following antibodies were used for immunohistochemistry: monoclonal mouse anti-human α-SMA (M0851, DAKO, Agilent Technologies, Inc., Santa Clara, CA, USA) and monoclonal mouse anti-human CD34 (M7165, DAKO, Agilent), both diluted 1:100. The detection system was EnVision™ FLEX+, Mouse, High pH (Link) (K8002, DAKO Cytomation, Agilent, Glostrup, Denmark). The manufacturer’s protocols were followed. Eighteen sections were used as controls.
Photomicrographs of representative immunohistochemical staining fields were obtained using an Olympus CX 21 microscope fitted with an Olympus C5050Z digital camera (Olympus Optical Co., Ltd., Tokyo, Japan).

Georgiev G.P., Yordanov Y., Olewnik Ł., Tubbs R.S., LaPrade R.F., Ananiev J., Slavchev S.A., Dimitrova I.N., Gaydarski L, & Landzhov B. (2024). Do the Differences in the Epiligament of the Proximal and Distal Parts of the Anterior Cruciate Ligament Explain Their Different Healing Capacities? Quantitative and Immunohistochemical Analysis of CD34 and α-SMA Expression in Relation to the Epiligament Theory. Biomedicines, 12(1), 156.

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Quantifying Angiogenesis in Cancer Models

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Example 7

Tissue sections were process as described (Sukhdeo, K. et al. Proc Natl Acad Sci USA 104, 7516-21 (2007)). Sections were incubated with primary antibodies (5 μg/ml) or the corresponding IgG fraction of preimmune serum overnight at 4° C. in blocking solution (3% BSA/PBS). BCL9 (ab37305, Abcam), mouse CD34 (RAM34, eBiosciences), human CD34 (M7165, Dako) and human CD44H (2C5, R&D Systems) antibodies were employed. Blood vessel formation in the CRC and MM models was evaluated using anti-mouse CD34 and anti-human CD34 antibodies, respectively, and the corresponding biotinylated antibodies coupled to streptavidin peroxidase (Vector). The number of blood vessels was determined by counting the mean number of independent blood vessels in 5 randomly selected fields at 50× magnification as highlighted by CD34 staining (brown color). See results in FIGS. 4b, 4e, 4i, 6.

US10703785B2. Targeting deregulated Wnt signaling in cancer using stabilized alpha-helices of BCL-9 (2020-07-07). None [US], None [US], None [US]. Inventors: Loren D. Walensky [US], Ruben Carrasco [US], Gregory H. Bird [US].

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Immunohistochemical Analysis of Blood Vessels

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Example 7

US11220532B2. Targeting deregulated Wnt signaling in cancer using stabilized alpha-helices of BCL-9 (2022-01-11). DANA-FARBER CANCER INSTITUTE, INC. [US]. Inventors: Loren D. Walensky [US], Ruben Carrasco [US], Gregory H. Bird [US].

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Immunofluorescence Staining of SFT/HPC Tissues

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Immunofluorescence staining was performed as described previously [24 (link)]. Briefly, FFPE tissues of SFT/HPC cases were cut into sections 3 mm thick and deparaffinized using xylene. Heat-induced antigen retrieval was performed in citrate buffer (pH = 6.0) for 15 min, and 3% hydrogen peroxide was applied to inactivate endogenous peroxidase activity. The sections were next incubated for 16 h at 4 °C with primary antibodies targeting CD34 (anti-mouse antibody, 1:50, M7165, DAKO) and CLDN5 (anti-rabbit antibody, 1:200, ab131259, Abcam). Subsequently, sections were co-incubated with goat anti-mouse IgG (1:200, A-11001, Life Technologies, Carlsbad, CA, USA) and goat anti-rabbit IgG (1:200, A1011, Life Technologies) secondary antibodies for 1 h at room temperature. Nuclei were counterstained with 300 nM 4′,6-diamidino-2-phenylindole (DAPI) for 20 min, followed by washing with PBS. The chamber slides were mounted with antifade mounting medium and imaged using the EVOS fluorescence imaging system (Thermo Fisher Scientific, Waltham, MA, USA).

Hong J.H., Noh M.G., Akanda M.R., Kim Y.J., Kim S.H., Jung T.Y., Jung S., Lee J.H., Rhee J.H., Kim K.K., Kim S.S., Lee K.H, & Moon K.S. (2021). Solitary Fibrous Tumor/Hemangiopericytoma Metastasizes Extracranially, Associated with Altered Expression of WNT5A and MMP9. Cancers, 13(5), 1142.

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Annotating Vascular Lakes in Uveal Melanoma

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All tumor specimens had been worked up for routine diagnostic histopathology, including staining for hematoxylin and eosin (H&E). Representative H&E-stained UM sections were scanned at 40× magnification using Aperio CS2 (Leica Biosystems, Newcastle-Upon-Tyne, UK) and saved as high-resolution WSIs. The annotation of the scanned WSIs of UM sections was performed using QuPath Bioimage analysis v 0.2.0-m8 (University of Edinburgh, Scotland, UK).¹⁸ (link) In some large tumors, several H&E sections were scanned; for comparison purposes, the tumor section with the largest number of VLs was assessed. H&E sections were removed from the cohort if they were hemorrhagic or necrotic, because of the difficulty to identify clear VLs.
For reproducibility, a VL was annotated only if it satisfied the following criteria:

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A VL must contain blood and/or plasma.

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Eighty percent or more of the “lake” must lack endothelial lining (Fig. 1).

Where it was uncertain whether a VL had an endothelial lining, immunostaining was undertaken using a CD34 antibody and red chromogen staining (M7165, dilution 1:50, clone: QBEND10; DAKO, Santa Clara, CA, USA).
Annotators were blinded from all patient data, including patient outcome, while the digital analysis was being performed (annotations completed by HJ and SEC).

Jones H., Kalirai H., Taktak A., Chen K, & Coupland S.E. (2022). Vascular Lakes in Uveal Melanoma and Their Association With Outcome. Translational Vision Science & Technology, 11(3), 32.

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Immunohistochemical Analysis of Tumor Samples

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Tumors were fixed in formalin and embedded in paraffin. Briefly, 5 μm-thick sections were cut using a microtome, transferred onto adhesive slides, and dried at 62°C for 30 min. Immunohistochemistry with antibodies against CD31 (M0823, Dako, CA, USA), CD34 (M7165, Dako, CA, USA), TFE3 (MRQ-37, CA, USA), and CAMTA1 (ab64119, Abcam, Cambridge, UK) was performed using an automated immunohistochemical staining instrument (Ventana Discovery^® XT, Ventana Medical System, AZ, USA).

Lee S.J., Yang W.I., Chung W.S, & Kim S.K. (2016). Epithelioid hemangioendotheliomas with TFE3 gene translocations are compossible with CAMTA1 gene rearrangements. Oncotarget, 7(7), 7480-7488.

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Immunohistochemical Staining for Cell Markers

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The immunohistochemical stain for CK7 (1 : 200, Dako M7018, mouse monoclonal, clone OV-TL 12/20), CK19 (1 : 50, Dako M0888, mouse monoclonal, clone RCK108), and CD34 (1 : 160, Dako M7165, mouse monoclonal, clone QBEnd 10) was performed on the 5-micrometer thick tissue sections of paraffin-embedded tissue blocks. Antigen retrieval was carried out with 0.01 M citrate buffer at pH 6.0. The slides were stained on the Dako Autostainer (Dako Corporation, Carpinteria, CA).
Sections of various control tissues (breast for CK7, liver for CK19, and colon for CD34) with known positivity for the target proteins were used as positive stain controls. Positive staining was defined as dark brown staining pattern. Scant or fine granular background staining or no staining at all was considered negative.

Lee H., Ainechi S., Dresser K, & Kurian E.M. (2014). Central Portalization Correlates with Fibrosis but Not with Risk Factors for Nonalcoholic Steatohepatitis in Steatotic Chronic Hepatitis C. International Journal of Hepatology, 2014, 329297.

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Gremlin-1 Immunohistochemistry in Aortic Tissue

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For immunohistochemical labelling, formalin-fixed paraffin-embedded aortic tissue sections of LDS patients or healthy donors were pre-treated in citrate buffer and incubated with antibodies against Gremlin-1 (bs-1475R, Bioss, Woburn, MA) in an automated stainer (Ventana Medical Systems, Tucson, AZ) according to a standard protocol (CC1st). For double labelling with Gremlin-1 and muscle actin (ENZ-30931, Enzo Life Sciences GmbH, Loerrach, Germany) or Gremlin-1 and CD34 (M7165, Dako, Hamburg, Germany), incubation with Gremlin-1 antibodies was followed by a short denaturing step and incubation with the second antibody. Bound antibodies were detected by the peroxidase method using diaminobenzidine as chromogen (760–500, Ultraview DAB, Ventana). For double labelling studies, bound Gremlin-1 antibodies were visualized with DAB as described above and expression of muscle actin or CD34 was demonstrated by alkaline phosphastase linked secondary antibodies using fast red as chromogen (760–501, Ultraview Universal Detection Kit, Ventana).

Wellbrock J., Sheikhzadeh S., Oliveira-Ferrer L., Stamm H., Hillebrand M., Keyser B., Klokow M., Vohwinkel G., Bonk V., Otto B., Streichert T., Balabanov S., Hagel C., Rybczynski M., Bentzien F., Bokemeyer C., von Kodolitsch Y, & Fiedler W. (2014). Overexpression of Gremlin-1 in Patients with Loeys-Dietz Syndrome: Implications on Pathophysiology and Early Disease Detection. PLoS ONE, 9(8), e104742.

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