For paraffin sections, whats better for decalcification of bones, I target the complex vertebra of catfish.
I would like to thank you twice, first for your swift response, second to direct me to this rich discussion, it is really helpful, but I still wonder, I would use the rapid decal methods (acids), they suggested formic acid, however the links to complete protocol not working. I read a method suggested nitric 5% for 3 days then benzene, is any acid useful in high concentration 5%, should I use benzene after acids, should other steps as dehydration, clearing will be performed, or embedding the samples in paraffin just after decal process.
When you mention the complex vertebra of a catfish, are you referring to the Weberian complex or to the complex vertebra in the caudal skeleton? What is the size (length versus width) of this vertebra? This information makes a lot of difference in the timing because if the vertebra is small only a few hours will be needed in the acid solution. The decalcification process is the beginning. I have had excellent results doing histological cross section of catfishes, just using the "old" method: after decalcification I washed the sample several times (about 12 hours in each) in ethanol 70, then ethanol 90, then ethanol 100 and then left the sample in benzene (or toluene) for about 12 hours. After this "washing process" you can embed the sample in paraffin and etc...
Thanks Dear Gloria, I target the Weberian complex, Its funnel shaped about 2.5-3 cm in length and about 1cm in diameter. For each sample I am looking for a rapid decal process, the suitable acid, the concentration %, the time allowed, and if any neutralization required or not. There are several choices, I could not decide what is the best ??
Dear Mohamed,
Is the vertebral centrum alone one cm in diameter? Because then this is a large vertebra that is probably heavily ossified. I used for large vertebrae a nitric solution for about 2 days, but I was constantly checking the bone to avoid over decalcification. You cannot have a fast process with such large bone. If you increase the concentration of the acid, then you may destroy the bone. For such size of a centrum you easily have to count about 3 to 5 days just decalcifying and washing the sample.
Dear Gloria
The centrum is hollow, the thickness is about one millimeter, I target the complex vertebra that lodged a part of the webberian ossicles (fused first to fifth vertebrae of clarias garipenus, standard length of fish about 30 cm). From your answers and other reports, I think that nitric acid 5% for 3 hours , no neutralization will be helpful, what is your opinion?
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