I'm conducting immunogold-staining with LR white.
Here is the protocol of dehydration
Conventional:
50% EtOH/50% water ? 15 min
70% EtOH/30% water ? 15 min
90% EtOH/10% water ? 10 min
100% EtOH ? 10 min
1:1 EtOH:Resin ? Overnight
100% Resin ? 2 hours
100% Resin ? 1 hour
For Immunocytochemistry:
Even though the tissue may be taken from 70% ethanol into LW White resin no special procedure is necessary. However, due to the fact that the nature of the tissue is not always consistent, we feel it is safer with at least one more change with 80% alcohol before going to resin.
50% EtOH/50% water ? 15 min
70% EtOH/30% water ? 15 min *
80% EtOH/20% Water 10 min
2:1 LR White resin to 70% EtOH 1 hour**
100% LR White resin 1 hour
100% LR White resin overnight
100% LR White resin 30 min
100% LR White resin 30 min
Why is 70% ethanol is used instead of 100% ethanol during dehydration process for immunocytochemistry?
Hello,
It is said in the document from EMS from which you cut and copied a part:
https://www.emsdiasum.com/microscopy/technical/datasheet/14380_LR_white.aspx
In conventional, samples have been post-fixed with osmium, making it stronger.
For immunocytochemistry, samples are not post-fixed with osmium to keep antibody epitopes. But you lose in strength. To avoid shrinkage, they recommend a step with water during embedding. That's the 2 asterisks referring to this remark.
It's not because of immunocytochemistry, but because LR white is water soluble and final steps of dehydration are performed with a resin, not with alcohol. You can see from your protocols that after full dehydration with alcohol only 2 changes of resin are needed, but after partial dehydration (up to 70% alcohol) 4 changes of resin are required.
Interesting ...
we can see that (not cited) C&P ='copying and pasting' induces sometimes also misunderstandings [as compared to the "Original" as found at:
https://www.emsdiasum.com/microscopy/technical/datasheet/14380_LR_white.aspx (eg: formation /display of ? in Jooeun Lee Jooeun Lee's 'Question')] :
("My"result for >Copy and paste< from there is:
>>Conventional:
50% EtOH/50% water - 15 min
70% EtOH/30% water - 15 min
90% EtOH/10% water - 10 min
....
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