H&E staining schedule

From:A van Binsbergen

    We have a Sakura DRS stainer (wonderful machine) and my staining
protocol is as follows:

3m sections - smalls stained immediately after cutting, large tissue - about
3mins draining before staining

dryer - about 65degrees celcius        5mins
xylene(dewax)     max agitation         1min 40sec
xylene                         "                    1min 30sec
absolute alcohol          "                    15sec
absolute alcohol          "                    15sec
wash (tap water)                              20sec
dist water                                         10sec
haematoxylin *            "                    3mins
wash (tap water)                              30sec
scotts (blueing agent)   "                   1min
wash (tap water)                              1min
eosin-phloxine #         "                    1min 20sec
wash (tap water)                              15sec
absolute alcohol          "                    15sec
absolute alcohol          "                    15sec
xylene                         "                    15sec
xylene                         "                    15sec

*mayers, homemade, double strength, changed every 2nd week(about 2000
# 2%eosin y, 1%phloxine B, aqueous, mixed  3:1, add 4drops conc. acetic
acid - changed every 2nd week

all solvents are rotated (see below) midweek and discarded at the end of
each week - all staining dishes are then soaked in detergent overnight,
rinsed and dried.

our staining is crisp and clear - no complaints from the paths.

protocol in use for more than 6months.

Rotation: dewax xylene discard, wipe dish, decant xylene 2 into dish
                wipe out xylene 2 dish and fill up with fresh xylene

or        remove dewax xylene, shift xylene 2 into that slot
            discard dewax and clean dish very well before refilling with
fresh xylene

do the same with the alcohols

each staining dish takes about 600ml of solvent so we are using about
4litres of xylene and 4litres of absolute alcohol each week

hope this helps

Annie in Africa

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