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    Fast Green/Sirius Red Protocol For Leica ST5020 Automated Stainer

    Zbigniew Mikulski, Angela Denn

    Published: 2024-06-08 DOI: 10.17504/protocols.io.ewov19djklr2/v1
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    Abstract

    This Sirius Red Fast Green protocol is for the Leica ST5020 Automated multistainer. You can use it as a guide to developing your own protocol, catered to the needs of your researchers. This template can be modified for manual staining but thebest and most reproducible results are easier to achieve with an automated stainer.

    This fork describes modifications to obtain high-quality special stain demonstrating collagen in FFPE and frozen tissues.

    Before start

    Steps

    Fast Green/Sirius Red Stain

    1.

    Perform all required maintenance on the stainer and prepare working solutions

    2.

    Arrange the reagents in the stainer according to the layout (note dark blue positions for FG/SR cups)

    Reagent map for ST5020
    Reagent map for ST5020
    3.

    Program the FG/SR protocol into the ST5020 stainer according to the table below

    Steps for staining
    Steps for staining

    Expected results

    4.

    Collagen fibers should be stained red and all other non-collagen proteins should be stained green.

    There is strong positive staining in collagen fibers along blood vessels and minimal red staining in other tissue areas. Normal mouse liver sample scanned with 40x 0.95NA objective with Zeiss AxioScan Z1
    There is strong positive staining in collagen fibers along blood vessels and minimal red staining in other tissue areas. Normal mouse liver sample scanned with 40x 0.95NA objective with Zeiss AxioScan Z1
    In this mouse tumor sample there are abundant collagen fibers stained red. Minimal background staining in other, non-collagenous areas of the tissue. Sample scanned with 40x 0.95NA objective with Zeiss AxioScan Z1
    In this mouse tumor sample there are abundant collagen fibers stained red. Minimal background staining in other, non-collagenous areas of the tissue. Sample scanned with 40x 0.95NA objective with Zeiss AxioScan Z1

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