Ar6 buffer

Manufactured by Akoya Biosciences

The AR6 buffer is a laboratory reagent used in various biological and biochemical applications. It is designed to maintain the proper pH and ionic conditions for the effective performance of specific experiments or assays. The core function of the AR6 buffer is to provide a controlled environment that supports the stability and activity of the target molecules or processes under investigation. No further interpretation or extrapolation on the intended use of this product is provided.

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Lab products found in correlation

Axio scan z1, by Zeiss (1 mentions) Decloacking chamber, by Biocare Medical (1 mentions) Opal 7 color manual ihc kit, by Akoya Biosciences (1 mentions) Protein block, by Agilent Technologies (1 mentions) Rabbit anti pd l1, by Cell Signaling Technology (1 mentions) Rabbit anti pd 1, by Cell Signaling Technology (1 mentions) Ar9 buffer, by Akoya Biosciences (1 mentions) Rabbit anti cd11b, by Cell Signaling Technology (1 mentions) Opal polymer hrp ms rb, by PerkinElmer (1 mentions) Opal 7 color manual ihc kit, by PerkinElmer (1 mentions) F6057, by Merck Group (1 mentions)

4 protocols using ar6 buffer

Multiplex Immunofluorescence Staining of Tissue Sections

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Tissue sections (5-μm) were baked at 60 °C for an hour and deparaffinized with xylene and a graded series of ethanol. Antigen retrieval was conducted using a Decloacking chamber (Biocare Medical) at 90 °C for 15 min in AR6 buffer (Akoya Biosciences). Multiplex fluorescent immunostaining was conducted following the Opal four-color user manual (Akoya Biosciences), including a nuclear 4′,6-diamidino-2-phenylindole (DAPI) counterstain. A tracheobronchial lymph node from a cat determined to no longer have systemic infection was used simultaneously as a negative control for GFP and a positive control for myeloid/histiocytic antigen and CD20. Whole-slide images were acquired using a Zeiss Axio Scan Z.1 whole-slide scanner at 200× equipped with a Colibri 7 LED light source and 16-bit Orcha Flash 4.0 monochrome camera. Immunohistochemical and acquisition parameters are outlined in SI Appendix, Table S3.

Nambulli S., Rennick L.J., Acciardo A.S., Tilston-Lunel N.L., Ho G., Crossland N.A., Hardcastle K., Nieto B., Bainbridge G., Williams T., Sharp C.R, & Duprex W.P. (2022). FeMV is a cathepsin-dependent unique morbillivirus infecting the kidneys of domestic cats. Proceedings of the National Academy of Sciences of the United States of America, 119(43), e2209405119.

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Multiplex Immunohistochemistry Staining Protocol

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mfIHC staining was performed using Opal 7-Color Manual IHC Kit (Akoya Biosciences), according to the manufacturer's recommendations. Concretely, 5 μm FFPE tissues were baked for 60 minutes at 60°C. After deparaffinization, a heat-mediated stripping procedure using AR6 buffer (Akoya Biosciences) at 95°C for 15 minutes was performed with the EZ-Retriever systems V.3 (BioGenex) between each Ab staining cycle. After blocking with Protein Block (Agilent Technologies Inc) for 20 minutes, slides were incubated for 30 minutes at room temperature with primary Abs. Sequentially, Opal Polymer HRP Ms + Rb was introduced for 10 minutes and preselected opal fluorophores were applied for 10 minutes at room temperature. The staining procedure was carried out in a step-by-step manner. After all staining was completed, slides were counterstained with 4′,6-diamidino-2-phenylindole (DAPI) solution (Supplementary Fig. S3A–S3D). Supplementary Table S1 contains the Abs and reagents for the four panels utilized.

Fukuda Y., Kim S.H., Bustos M.A., Cho S.N., Roszik J., Burks J.K., Kim H., Hoon D.S., Grimm E.A, & Ekmekcioglu S. (2023). Inhibition of Microsomal Prostaglandin E2 Synthase Reduces Collagen Deposition in Melanoma Tumors and May Improve Immunotherapy Efficacy by Reducing T-cell Exhaustion. Cancer Research Communications, 3(7), 1397-1408.

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Multiplex Immunofluorescence Tissue Staining

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Slides were deparaffinised in Xylene and rehydrated in a series of ethanol dilutions. Using the Leica Bond automated staining robot, samples underwent heat-induced antigen retrieval (HIER) of 30 min at 100 °C. Then tissue slides were exposed to multiple staining cycles each including a 30 min incubation with a protein block (Akoya), 1 h incubation with the respective primary antibody, 30 min incubation with the secondary antibody (Akoya), 10 min incubation with the respective OPAL (Akoya) followed by 20 min incubation with AR6 buffer (Akoya) at 85 °C prior to the next staining cycles and finally stained with fluorescent DAPI (Akoya) for 10 min. In between each step, slides were washed with bond wash for 5 min.
Primary antibody concentrations and OPAL pairings are shown in Supplementary Table S2. Antibody-OPAL pairings were assigned based on expected biomarker abundance and expected co-expression. Dilution of antibodies was assessed by single stains.

Mathieson L., Koppensteiner L., Dorward D.A., O’Connor R.A, & Akram A.R. (2024). Cancer-associated fibroblasts expressing fibroblast activation protein and podoplanin in non-small cell lung cancer predict poor clinical outcome. British Journal of Cancer, 130(11), 1758-1769.

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Multiplex Immunofluorescence Protocol for DLBCL TMA Analysis

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The DLBCL TMA slides were stained with multiplex uorescence by using the Opal 7-color Manual IHC Kit (PerkinElmer, MA). After dewaxing by xylene and rehydration by ethanol, slides were heated in a microwave with AR6 Buffer (AR600, AKOYA) and AR9 Buffer (AR900, AKOYA) for antigen retrieval. The slides were incubated with primary antibodies overnight at 4°C and then incubated with secondary antibody for 10min at room temperature. At last, we used 4',6-diamidino-2-phenylindole (DAPI; F6057, Sigma) to stain the nuclei and seal the slides. Imaging was achieved using the Vectra 3.0 Automated Quantitative Pathology Imaging System. Tumor and stroma images were captured at ×20 magni cation. Finally, the staining was scored by inForm® Cell Analysis software based on the intensity and degree of staining.
The primary antibodies used in this study were as follows: rabbit anti-PHKA1 (24279-1-AP, Proteintech), rabbit anti-PLTP (ab282456, Abcam), rabbit anti-CD163 (93498, Cell Signaling Technology), rabbit anti-CD68 (76437, Cell Signaling Technology), rabbit anti-CD11B (49420, Cell Signaling Technology), mouse anti-CD66b (ARG66287, Arigobio), rabbit anti-PD-1 (86163, Cell Signaling Technology) and rabbit anti-PD-L1 (13684, Cell Signaling Technology). The secondary antibody was Opal™ polymer HRP Ms+Rb (ARH1001EA, Perkin Elmer).

He J., Chen Z., Xue Q., Sun P., Wang Y., Zhu C., & Shi W. (2022). Identification of molecular subtypes and a novel prognostic model of diffuse large B-cell lymphoma based on a metabolism-associated gene signature.

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