Multiplex IHC was performed as previously described (34 (link)), using Opal Automation IHC Kit (Akoya Bioscience) and Bond RXm autostainer (Leica Biosystems). The following antibodies were used: anti-pan cytokeratin (rabbit poly; RRID: AB_10855057, Bioss), anti-PDPN (clone 66; RRID: AB_2785565, Invitrogen), anti-αSMA (clone EPR5368; RRID: AB_11129103, Abcam), anti-CD31 (clone D8V9E; RRID: AB_2722705, Cell Signaling Technology), anti-VEGFR3 (clone AFL4; RRID: AB_467795, Thermo Fisher Scientific), and anti-digoxigenin (DIG, clone 9H27L19; RRID: AB_2532342, Thermo Fisher Scientific; 1:500). Cells were counterstained with DAPI. Stained slides were imaged with Mantra Quantitative Pathology Workstation (Akoya Biosystems), and images were analyzed with inForm software (RRID: SCR_019155, Akoya Biosystems).
Opal automation ihc kit
Manufactured by Akoya Biosciences
The Opal Automation IHC Kit is a laboratory equipment product designed for automated immunohistochemistry (IHC) processing. It provides a standardized and consistent approach to performing IHC staining procedures. The core function of the kit is to enable automated, reproducible, and high-throughput IHC analysis of tissue samples.
Automatically generated - may contain errors
Lab products found in correlation
Bond rxm autostainer, by Leica (2 mentions)
Anti pan cytokeratin rabbit poly, by Bioss Antibodies (1 mentions)
Bond polymer refine red detection kit, by Leica (1 mentions)
Vs120, by Olympus (1 mentions)
E cadherin, by Cell Signaling Technology (1 mentions)
Prolong diamond, by Thermo Fisher Scientific (1 mentions)
Anti ki 67 clone d3b5, by Cell Signaling Technology (1 mentions)
Anti dig clone 9h27l19, by Thermo Fisher Scientific (1 mentions)
3 protocols using opal automation ihc kit
1
Multiplex IHC for Tissue Characterization
2
Multiplex ISH and E-cadherin IF for Mouse Rest and Chga
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Chromogenic In situ hybridization (ISH) for mouse Rest (RNAscope® LS 2.5 probe–Mm-Rest, ACDBio #316258) or mouse Chga (RNAscope® LS 2.5 probe–Mm-Chga, ACDBio #447858) was performed on a Leica Bond RX with Bond Polymer Refine Red Detection kit DS9390). For multiplex ISH and E-cadherin immunofluorescence (IF), ISH was detected with Opal650 using the Opal automation IHC kit (Akoya Biosciences, SKU NEL821001KT), followed by E-cadherin (Cell Signaling Technology 3195, 1:400) on the Leica Bond Max with EnVision+ System-HRP Labeled Polymer Anti-Rabbit (DAKO, K4003) and Opal570 followed with DAPI. Whole sections were scanned using the Olympus VS120 and staining was quantified using the ISH detection algorithms in HALO.
3
Multiplex IHC for Tumor Cell Profiling
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Multiplex immunohistochemistry was carried out as described previously
18 (link) using an Opal Automation IHC Kit (Akoya Bioscience) and Bond RXm autostainer (Leica Biosystems). The sections were stained with DAPI and the following Abs: anti‐CD45 (clone D3F8Q; Cell Signaling Technology; 1:500), anti‐DIG (clone 9H27L19; Thermo Fisher Scientific; 1:500), anti‐Ki‐67 (clone D3B5; Cell Signaling Technology; 1:500), and anti‐pCK (rabbit poly; Bioss Antibodies; 1:250). Stained slides were mounted with ProLong Diamond (Thermo Fisher Scientific) and imaged with a Mantra Quantitative Pathology Workstation (Akoya Biosystems). The obtained images were analyzed with inForm Tissue Finder software (Akoya Biosystems). To calculate the percentage of Ki‐67 positive cancer cells, inForm software was trained to detect tissue and cell phenotypes using machine‐learning algorithms based on the following criteria: areas with pCK expression = tumor, other areas = stroma, pCK+ CD45− cells = cancer cells, pCK− CD45+ = blood cells, and pCK− CD45− = other cells. inForm software computed the percentage of Ki‐67‐positive cells among cancer cells. The average percentage was calculated from five images for each specimen.
18 (link) using an Opal Automation IHC Kit (Akoya Bioscience) and Bond RXm autostainer (Leica Biosystems). The sections were stained with DAPI and the following Abs: anti‐CD45 (clone D3F8Q; Cell Signaling Technology; 1:500), anti‐DIG (clone 9H27L19; Thermo Fisher Scientific; 1:500), anti‐Ki‐67 (clone D3B5; Cell Signaling Technology; 1:500), and anti‐pCK (rabbit poly; Bioss Antibodies; 1:250). Stained slides were mounted with ProLong Diamond (Thermo Fisher Scientific) and imaged with a Mantra Quantitative Pathology Workstation (Akoya Biosystems). The obtained images were analyzed with inForm Tissue Finder software (Akoya Biosystems). To calculate the percentage of Ki‐67 positive cancer cells, inForm software was trained to detect tissue and cell phenotypes using machine‐learning algorithms based on the following criteria: areas with pCK expression = tumor, other areas = stroma, pCK+ CD45− cells = cancer cells, pCK− CD45+ = blood cells, and pCK− CD45− = other cells. inForm software computed the percentage of Ki‐67‐positive cells among cancer cells. The average percentage was calculated from five images for each specimen.
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!