Automated immunohistochemical stainer

Manufactured by Agilent Technologies

The Automated Immunohistochemical Stainer is a laboratory instrument designed to automate the process of immunohistochemical staining. It performs the critical steps required for this technique, including sample preparation, reagent application, and incubation, in a controlled and standardized manner.

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Lab products found in correlation

Citrate buffer, by Agilent Technologies (1 mentions) Envision dual link system, by Agilent Technologies (1 mentions) Envision detection system, by Agilent Technologies (1 mentions)

2 protocols using automated immunohistochemical stainer

Immunohistochemical Analysis of YAP and AREG

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Slides from Tumor paraffin‐embedded blocks were pretreated with 0.01 mol/L citrate buffer (pH 6; Dako) for 20 minutes at 100°C, then immunostainings were performed with automated immunohistochemical stainer (Dako). Slides were successively incubated at room temperature in 3% H₂O₂ for 5 minutes, then with monoclonal antibody (Table S4) diluted at 1:200 for 60 minutes at room temperature. Finally, antibody fixation was revealed by the EnVision+ Dual Link System (Dako).
The staining intensity was recorded by two expert pathologists (MA and CD) blinded to treatment arm. YAP was scored as 0 (negative), 1 (weak), 2 (moderate) or 3 (strong), at ×40 magnification. An overall IHC composite score was calculated from the sum of the staining intensity (0‐3) multiplied by the distribution (0%‐100%) from all parts of the slide, giving a H‐score between 0 and 300. AREG was scored as negative (no signal despite positive internal control) or positive at ×40 magnification.

Maille E., Levallet J., Dubois F., Antoine M., Danel C., Creveuil C., Mazieres J., Margery J., Greillier L., Gounant V., Moro‐Sibilot D., Molinier O., Léna H., Monnet I., Bergot E., Langlais A., Morin F., Scherpereel A., Zalcman G, & Levallet G. (2022). A defect of amphiregulin release predicted longer survival independently of YAP expression in patients with pleural mesothelioma in the IFCT‐0701 MAPS phase 3 trial. International Journal of Cancer, 150(11), 1889-1904.

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Quantifying VPS4 Protein Levels in CRC

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The study protocol for analysis of protein levels of VPS4B and VPS4A in human normal colon and CRC samples was approved by the Bioethics Committee of the Maria Skłodowska‐Curie Institute‐Oncology Centre in Warsaw (decision no. 40/2017). The informed consent was obtained from all subjects. The experiments conformed to the principles set out in the WMA Declaration of Helsinki and the Department of Health and Human Services Belmont Report. High‐density tissue microarrays were constructed from formalin‐fixed, paraffin‐embedded diagnostic samples of 100 pairs of treatment‐naïve CRC tissues, and matched normal colon samples from the collection of the Maria Skłodowska‐Curie Institute‐Oncology Centre. IHC was performed using automated immunohistochemical stainer (Dako Denmark A/S) and anti‐VPS4B or anti‐VPS4A antibodies (details above). The EnVision Detection System (Agilent) was used for detection. Samples were reviewed for abundance of VPS4 proteins in normal and neoplastic tissues by two pathologists, who were blinded to outcome. A semi‐quantitative method was applied for IHC evaluation, involving a scoring system based on the staining intensity: 0—no staining; 1—weak, 2—intermediate, and 3—strong staining; staining homogeneity was above 90%.

Szymańska E., Nowak P., Kolmus K., Cybulska M., Goryca K., Derezińska‐Wołek E., Szumera‐Ciećkiewicz A., Brewińska‐Olchowik M., Grochowska A., Piwocka K., Prochorec‐Sobieszek M., Mikula M, & Miączyńska M. (2020). Synthetic lethality between VPS4A and VPS4B triggers an inflammatory response in colorectal cancer. EMBO Molecular Medicine, 12(2), e10812.

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