[Histonet] Mouse Tissue Processing Problems Help!

djemge <@t> aol.com djemge <@t> aol.com
Thu Oct 27 10:46:39 CDT 2005

Please help! Most of the mouse tissue (liver, brain, embryo's, pancreas) is very dry at times almost glassy. I came from a clinical histology lab and recently accepted a position as the only histotech in a research lab. The main tissue in this lab is Bouins fixed gonadal mouse tissue rinsed with water for 3 days, then 50% ETOH for 3 days, and 4 to 8 hour BNF fixed mouse tissue that is transferred to 70% ETOH (For later IF and IHC) then given to me. The Bouins fixed mouse testes and epididymous seem fine although somewhat crystal like at the inside edges. Formalin fixed testes transferred to the 70% has no crystals but is dry. All parameters and protocols were established by my predecessor. Since I am not familiar with mouse tissue I want to get this right!
Processor Set up on Sakura VIP 2000 Enclosed Processor
Using Surgipath Reagent Alcohol
70% Alcohol             30 mins    Retort temp 38 degees C   P/V on  (the tissue sits in this for delay)
70% Alcohol             30 mins                      "                                "
80% Alcohol             30 mins                       "                               "
95% Alcohol             30 mins                       "                               "
100% Alcohol           30 mins                      "                                "
100% Alcohol           30 mins                       "                               "
100% Alcohol            30mins                       "                               "
Xylene                        30mins                       "                                "
Xylene                         30mins                      "                                "
Xylene                         30mins                      "                                "
Paraffin                       30mins        Temp set at 58 degrees C  (although retort temp says 60 C)  P/V  on  
Paraffin                       30mins                            "                                                                                      "     
Paraffin                       30mins                            "                                                                                      "
Help with Paraffin: 
The Paraffin was Surgipath Formula R, but this crackled when put on ice. I switched to Cardinal Health's Ameraffin Cat no. M7346-1A I changed the processor station temps and embedding station temps to 58 degrees C per manufactures instructions. I can't say I am satisfied with this paraffin either as it is sticky and I need a waterbath temp of 38 degrees to cut it or it spreads quickly (I used this at my old lab for human tissue, but it seems to respond differently here - I manually checked the temps). The Formula R had a waterbath temp of 47 degrees and my predecessor had the processor station temps at 60 C and the retort temp had 63 to 64 C indicated, and embedding center at 62C. 

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