Hello Agree with comments about temperature of 60 degrees killing an enzyme. If you plot enzyme activity on "y" axis and temperature on "x" axis it is not a straight line. Every enzyme has an optimal temperature and can function slowly at non-optimal or optimally at correct temperature. Diastase/amylase from your body work at 37 degrees, less so or dead at RT or 60 degrees. Nuking a mixture to 37 degrees might nuke protein. Nuke buffer to 37, verify temp and add the enzyme. pH-if you plot enzyme activity on "y" axis and pH on x-axis, it is not a straight line. There is an optimal pH for each enzyme. The enzyme might work fine at different pH; just not nearly as well. I know some still use 0.5% or 1.0% in di water. I'd advise not to. Use a buffer at a given pH. "pHíng" di or distilled water with ordinary lab electrodes is an act of futility. From a standard lab electrode point of view, there is NO pH to read. Not enough ions present so meter might say anything. If anything, it will pick up CO2 from air which becomes carbonic acid and meter will flail wildly in acidic region. But that is not a stable buffer. di water might work. Just that sometimes, it might not. Why chance it? di water has unstable (no reliably readable) pH. Ray Koelling, golfing in Spokane WA and enjoying the low humidity
----- Original Message ----- From: "Joanne Clark via Histonet" <histonet@lists.utsouthwestern.edu> To: histonet@lists.utsouthwestern.edu Sent: Wednesday, May 4, 2016 1:02:48 PM Subject: [Histonet] PAS Stain Hi Histonetters, I'm hoping someone can help me troubleshoot our PAS diastase method. We have been digesting the tissue in 0.5% diastase of malt in a 60 degree oven for 30 minutes, but do not see the glycogen being digested out. I have tried alpha amylase and beta amylase also without any luck. Does anyone have any suggestions to get the digestion to work Joanne Clark, BAAS, HT(ASCP)CM Director of Histology P. (575) 622-5600 C. (575) 317-6403 F. (575) 622-3720 TF. (800) 753-7284 pcnm.com Disclaimer: This electronic message may contain information that is proprietary, confidential, or legally privileged or protected. It is intended only for the use of the individual(s) and entity named in the message. If you are not an intended recipient of this message, please notify the sender immediately and delete the material from your computer. Do not deliver, distribute or copy this message and do not disclose its contents or take any action in reliance on the information it contains. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
