Dewaxing and pre-treatment was done as described above. Sequential double immuno-staining was performed using anti-human K10 clone DE-K10 (1:50) in combination with anti-human CD14 clone SP53 (1:200). In the first sequence, slides were incubated with K10 for 30 min (32 °C) and reactions were visualized using EnVision™ FLEX /HRP/DAB Detection Reagent (Agilent/Dako, Cat. no. GV800 + 821) following the recommendation of the manufacturer. After the visualization with DAB, slides were subsequently incubated with K14 for 30 min (32 °C). The reactions for K14 were detected with EnVision™ FLEX /HRP Detection Reagent (Agilent/Dako, Cat. no. GV800 + 821) and visualized with Magenta Substrate chromogen system (Agilent/Dako, Cat. no. GV925) following instructions given by the vendor. All slides, both single and double stained, were counterstained with Haematoxylin and mounted with Pertex.
Magenta substrate chromogen system
Manufactured by Agilent Technologies
The Magenta Substrate chromogen system is a laboratory equipment product designed to be used in immunohistochemistry and other biological applications. It functions as a chromogen, providing a visible color reaction when combined with specific target analytes. The core purpose of this product is to enable the visualization and detection of target molecules within a sample.
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Lab products found in correlation
Renoir red, by Biocare Medical (1 mentions)
Envision flex hrp reagent, by Agilent Technologies (1 mentions)
Envision flex antibody diluent, by Agilent Technologies (1 mentions)
Cd20 clone l26, by Agilent Technologies (1 mentions)
Envision flex target retrieval solution 3 in 1 ph 9, by Agilent Technologies (1 mentions)
2 protocols using magenta substrate chromogen system
1
Sequential Double Immunostaining of K10 and CD14
2
Dual Immunolabeling of PAX5 and CD20
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Double-immuno-labelling experiments were performed on 3-µmthick FFPE sections, using anti-human PAX5 clone DAK-Pax5 (Agilent/Dako, cat. no. M7307, Glostrup, Denmark) and CD20 clone L26 (Agilent/Dako, cat. no. M0755). All incubation steps were performed automatically on the Omnis (Agilent/Dako). Briefly, FFPE sections were exposed to deparaffinization, followed by antigen retrieval using EnVision™ FLEX Target Retrieval solution (3-in-1) pH 9 (Agilent/Dako, cat. no. GV800/821) at 97°C (24 min). In the first sequence, slides were incubated with PAX5 diluted 1:30 in Renoir Red (Biocare Medical, cat. no. PD904, Pacheco, CA, USA) for 30 min (32°C), and reactions were visualized with EnVision™ FLEX /HRP/DAB Detection Reagent (Dako, cat. no. GV800/GV821). After visualization with DAB, slides were incubated with CD20, diluted 1:500 in Envision Flex Antibody Diluent (Agilent/Dako, cat. no. K8006) for 30 min (32°C), reactions were detected with EnVision™ FLEX/HRP Reagent (Dako, cat. no. GV800/GV821) and visualized using Magenta Substrate chromogen system (Agilent/Dako, cat. no. GV925). All slides were counterstained with haematoxylin and mounted with Pertex.
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