S236784 2

Manufactured by Agilent Technologies

The S236784-2 is a laboratory equipment product manufactured by Agilent Technologies. It serves a core function, but a detailed description while maintaining an unbiased and factual approach is not available.

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Lab products found in correlation

Dulbecco phosphate buffered saline (dpbs), by Thermo Fisher Scientific (2 mentions) Superfrost plus adhesion slides, by Thermo Fisher Scientific (1 mentions) Anti cd68, by Cell Signaling Technology (1 mentions) Mp 7451, by Vector Laboratories (1 mentions) Epr3864, by Abcam (1 mentions) Sm803, by Agilent Technologies (1 mentions) Bp39920, by Thermo Fisher Scientific (1 mentions) Slf0601, by Bio-Rad (1 mentions) Decloaking chamber, by Biocare Medical (1 mentions)

3 protocols using s236784 2

SARS-CoV-2 Spike and Nucleocapsid Protein Staining

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5μm- sections of FFPE tissues on SuperFrost Plus adhesion slides (ThermoScientific, Cat. 10149870) were deparaffinized using 3 solutions of absolute m-xylene or Histoclear II histology (SLS, Cat. NAT1334), rehydrate in 90%, 80%, 70% ethanol solutions and DPBS (ThermoFisher, Cat.14190169). After then, the slides were immersed in an antigen retrieval solution pH 9.0 (Agilent Dako, S236784-2) and kept in a pressure cooker for 2 minutes. The cells into tissues were permeabilized in a 0.1%Tween20 solution for 5 minutes. The blocking, staining with primary and secondary antibodies were realized according to the manufactures of the following IHC kits: Mouse and rabbit specific HRP/DAB IHC detection Kit micro-polymer (Abcam, Cat.ab236466) and ImmPRESS[R] Duet double staining kit anti-rabbit AP/anti-mouse HRP (Vector laboratories, Cat.mp7724). Anti-SARS-CoV-2 spike antibody, targeting the S2 subunit, and anti-SARS-CoV-2 nucleocapsid (1:300) antibody were acquired from Insight Biotechnology (Cat. GTX632604) and BioserverUK (Cat.BSV-COV-AB-13), respectively. Anti-CD68 was obtained from Cell signalling (Cat. 7643T). All samples were stained with haematoxylin.

Trevelin S.C., Pickering S., Todd K., Bishop C., Pitcher M., Garrido Mesa J., Montorsi L., Spada F., Petrov N., Green A., Shankar-Hari M., Neil S.J, & Spencer J. (2022). Disrupted Peyer’s Patch Microanatomy in COVID-19 Including Germinal Centre Atrophy Independent of Local Virus. Frontiers in Immunology, 13, 838328.

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Quantifying ERG Expression in Tumor Samples

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Tumors from different treatment conditions (±HS, HSP70^−/−) were harvested 48 hours after treatment with 5-FU or PBS as a control. These tumors were placed in tissue storage (Miltenyi, 130-100-008) and then left for a minimum of 48 hours in a formalin bath to fix the tissues. After this step, tumors were included in paraffin and were then cut into 4 µm thick slices. The paraffin was then removed using the PT-link device (Agilent S236784‐2). Slides were labeled for 1 hour with the anti-ERG monoclonal antibody (1/200; Abcam, EPR3864), then stained with ImmPRESS HRP Goat Anti-Rabbit IgG (Peroxidase) for 30 min (Vector, MP-7451) to amplify the signal. Finally, slides were stained with DAB (Agilent, SM803 and DM827) and counterstained with hematoxylin (Enzo, ENZ‐ACC106) and were mounted and digitally scanned using the Nanozoomer HT2.0 device (Hamamatsu). Staining quantification was carried out using QuPath software (v.01.3 ref). The appropriate threshold to discriminate positive cells from negative was determined on positive controls (endothelial cells of large vessels). To analyze the whole cohort, we generated two scripts. The first one was dedicated to upload 12 rectangles of 160,000 µm². These areas were then randomly placed on slides by the investigator. The second script detected ERG-positive cells in those areas. Then the percentage of positive cells was calculated.

Pilot T., Fratti A., Thinselin C., Perrichet A., Demontoux L., Limagne E., Derangère V., Ilie A., Ndiaye M., Jacquin E., Garrido C., Ghiringhelli F., Chalmin F, & Rébé C. (2020). Heat shock and HSP70 regulate 5-FU-mediated caspase-1 activation in myeloid-derived suppressor cells and tumor growth in mice. Journal for Immunotherapy of Cancer, 8(1), e000478.

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Immunohistochemistry in FFPE Tissue

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Formalin-fixed paraffin-embedded (FFPE) human tissues were received mounted on adhesive slides. The slides were baked overnight in an oven at 55°C (Robbin Scientific, Model 1000) and an additional 30 minutes at 65°C (Clinical Scientific Equipment, NO. 100). Tissues were deparaffinized with xylene and rehydrated with graded ethanol baths. Two step antigen retrieval was performed in the Decloaking Chamber (Biocare Medical) using the following settings: set point 1 (SP1), 125°C, 30 seconds; SP2: 90°C, 30 seconds; SP limit: 10°C. Slides were further incubated in hot Target Retrieval Solution, pH 9 (Agilent, S236784–2) for 15 minutes. Slides were then washed in two brief changes of diH2O (~2 seconds) and once for 5 minutes in 1x phosphate buffered saline (PBS), pH 7.4 (Fisher, BP39920). Sections were blocked in 10% normal goat serum (NGS, Vector S-1000), 1% bovine serum albumin (BSA, Sigma A7906) in PBS for 30 minutes at 20°C in a humid chamber, followed by PBS washes. Primary antibodies were diluted in 5% NGS, 1% BSA in 1x PBS and applied overnight at 4°C in a humid chamber, covered with plastic coverslips (Bio-Rad, SLF0601). Following overnight incubation, tissues were washed 3 × 10 min in 1x PBS. Coverslips (Corning; 2980–243 or 2980–245) were mounted in Slowfade Gold plus DAPI mounting media (Life Technologies, S36938).

Burlingame E.A., Eng J., Thibault G., Chin K., Gray J.W, & Chang Y.H. (2021). Toward reproducible, scalable, and robust data analysis across multiplex tissue imaging platforms. Cell Reports Methods, 1(4), 100053.

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