Re: [Histonet] non-xylene processing
ewj <@t> pigsqq.org
ewj <@t> pigsqq.org
Wed Jul 17 16:11:35 CDT 2013
We used Dawn until we ran out. Dawn can work. It is hard to get here.
We used calgon with the dawn because the water is so hard.
> -------Original Message-------
> From: Michelle Moore <mmooreht <@t> yahoo.com>
> To: ewj <@t> pigsqq.org <ewj <@t> pigsqq.org>, histonet <@t> lists.utsouthwestern.edu <histonet <@t> lists.utsouthwestern.edu>
> Subject: Re: [Histonet] non-xylene processing
> Sent: Jul 18 '13 03:27
> We also de-wax slides in a similar way as the procedure listed below. We
> also use a waterbath and dish soap (Dawn)/rinse agent method. (Rinse agent
> (Cascade) helps with water tension.) The key is water temp. it has to be
> over 90 C in order for it to work. Obviously if you de-wax non-chemical you
> do not have the alcohols on the stain line either = huge cost savings all
> the way around, not to mention the healthier work area!
> Michelle Moore
> St. Thomas, USVI
> FROM: "ewj <@t> pigsqq.org" <ewj <@t> pigsqq.org>
> TO: histonet <@t> lists.utsouthwestern.edu
> SENT: Wednesday, July 17, 2013 1:01 PM
> SUBJECT: [Histonet] non-xylene processing
> We now have a no-xylene method start to finish
> We have a Midea (Mei Di) 800W manual dial microwave oven
> that we have rigged with a K-type metal shielded thermocouple
> that goes in through a slightly enlarged vent hole in the internal
> guide housing. The internal length of the thermocouple wire is
> wrapped with heavy duty aluminum foil to block microwaves from
> creating silly currents in the thermocouple wire. PT100 RTD's
> are great for water baths but microwaves zap them in seconds.
> The hysteresis temp controller switches a 12v relay which drives
> a 10A 240V relay that supplies power to a plain socket for the microwave
> The microwave oven is a manual dial type so just cutting the power
> turns it off and on quite effectively. A low power (% time) setting
> is enough. High settings give too much overshoot.
> The 12V DC circuit allows us to eliminate problems with the
> back EMF from the MW oven and 240V relay. We
> have a diode that blocks the DC back EMF.
> Cut in well fixed tissues - fixed in formalin or Bouins.
> Microwave Cassettes in 100% Isopropanol at 60C - 10 min
> Repeat with fresh IsopOH another 10 min.
> Put cassettes in a fired clay earthenware pot -
> Pour in paraffin heated to 75C in a metal teapot.
> Microwave 10 min at 80C
> Pour out paraffin into open metal pot.
> Repeat above 10 min.
> Pour out paraffin to metal pot.
> Take cassettes to the embedding station,
> Embed, ice, trim, section.
> Bake 20 min on slide warmer at 60C
> We dewax with 95C tap water heated in a tank.
> A PT100 RTD drives a hysteresis controller.
> Also we use 12 V relay switched by the controller to drive the 240V
> relay that switches the heating element on and off.
> We have an electric solenoid valve and a waterproof switch to control the
> water flow when we need the hot water.
> The hot water flows into a 1 L pot with silicone rubber handles.
> We add 20 gm of a dishwasher detergent powder made in Shanghai.
> (It's similar to Cascade. We dont like Finish very much as sold in China).
> We put in the slide racks after the water and detergent are in the pot.
> We treat for one minute, remove the slides, pour out the
> water and repeat for another 1 minute soak.
> Next comes a 30 sec soak in clear hot tap water
> and a 1 min rinse in plain running tap water.
> 0.5% Periodic acid 10 min
> Tap water rinse 1 min
> Distilled water 1 min
> Harris Hematoxylin 4 min
> Rinse tap water 1 min
> 1% HCl in 70% EtOH 30 sec
> Rinse and blue in tap water 1 min
> Eosin/Biebrich Scarlet 30 sec
> Rinse tap water 1 min
> Rinse Distilled Water 30 sec
> Dry with hair dryer
> Our mounting medium contains xylene as a clearing agent/diluent.
> That's the only xylene in the normal runs.
> We like this method because it is quick, environmentally friendly, and
> people friendly
> and it produces very lovely slides that we can read and photograph for our
> A major part of our work is diagnosis of pig diseases, so the ability to
> give same-day
> histopath results on formalin fixed samples is valuable to our customers.
> We do still use xylene to dewax formalin fixed paraffin block sections that
> are used for PCR presently,
> but we think we can use hot soapy water for that also with some larger
> We heat the teapot on an induction stove. We have that rigged for thermal
> control also
> and a PT100 in through the lid in the teapot. It is much trickier
> controlling the induction
> stove, because they have many built-in safety features and you can't just
> turn one off and
> on again at the mains outlet. What you have to do is solder in leads to a
> relay across the
> on-off push button, and rig the relay for momentary contact by including a
> capacitor that is charged
> for "on" and discharged for "off", with a momentary contact at each time
> faking a key press.
> This lets us heat our paraffin teapot accurately with induction stove
> efficiency and automatic temp control.
> E. Wayne Johnson
> Enable Ag Tech
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