Alcian Blue/PAS stain
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This is a combined method utilizing the properties of both the PAS (see PAS) and Alcian blue (see Alcian blue) methods to demonstrate the full complement of tissue proteoglycans.
The rationale of the technique is that by first staining all the acidic mucins with Alcian blue, those remaining acidic mucins which are also PAS positive will be chemically blocked and will not react further during the technique. Those neutral mucins which are solely PAS positive will subsequently be demonstrated in a contrasting manner. Where mixtures occur, the resultant color will depend upon the dominant moiety.
The combined result from the port-manteaux method demonstrates both acidic- neutral- and mixtures of acidic and neutral mucins.
1) Bring sections to distilled water.
2) Stain with Alcian blue 15 mins
3) Wash well in running tap water 2 mins
4) Rinse in distilled water
5) Treat with periodic acid 5 mins
6) Wash well in distilled water
7) Stain with Schiff's reagent 10 mins
8) Wash well in running tap water 5 mins
9) Stain nuclei with hematoxylin 1 min
10) Wash in running tap water 2 mins
11) Differentiate with acid alcohol
12) Wash and blue nuclei in Scott’s tap water
13) Wash in water
14) Dehydrate, clear and mount
acidic mucins: blue
neutral mucins: magenta
mixtures of above: blue/purple
nuclei: deep blue