Dear researchers
I have some samples from the stems of watermelon plants and I want to fix them in FAA fixative solution for about 6 months; then I will dehydrate, embed, and stain them for histology analysis. Is it possible to maintain samples for about 6 month in FAA fixative solution?
If not, how can I maintain them for 6 months or even more?
thank you
#FAA #histology #embedding #fixation #staining # Formaldehyde_Acetic_Acid
Put the samples in a fridge. At 4 degrees C samples can stay up to a year and still show good morphology; at least the samples I had did (tube worm samples from deep water hot vents).
Brent E Gowen Dear Thank you for your answer. It much appreciated!
OK, I will put my sample in a refrigerator.
Thank you so much.
hello moien
put your samples in fridge and check them every three days.
Thank you for your response dear Hamidreza Soufi
as Brent E Gowen said, I fixed them in FAA, then I put them in a fridge. However, I wonder why should I check them every three days? I mean which characteristics should I check in my samples twice a week???
Your samples were stem? or another part of plant?
Yup, I second that. What should you be looking for every 3 days ?
The sample may go brown as chlorophyll is extracted; but this usually happens when plants material is exposed to alcohols; the chlorophylls dissolve in alcohols. If left in FAA, minimal chlorophyll extraction should occur, especially at 4 degrees Celsius.
FAA can change its pH over time while interacting with a sample which can change the morphological appearance of a sample, but again, 4 degrees C helps to minimise this.
The fact that you are looking at plant material is also a bonus; the cell walls and morphological substructure is much more robust than in animal material.
If you are looking for gross histology sections in paraffin etc; your samples should look fine, even after 6 months. If you are interested in immunolabeling or specific histochemical staining, 6 months in FAA is not ideal.
Cheers.
B
Dear Brent E Gowen
Thank you for your great answer. I appreciate it.
Actually my field of research is #vegetable_grafting.
My samples are the grafting zone (graft union) of watermelon grafted unto Cucurbit as a rootstock.
I want to check Callus formation and vascular connection between scion and rootstock during the days after grafting in each treatment.
therefore, I should embed them in paraffin, as you mentioned, or GMA for histological analysis.
Best
Moein
In general, a disadvantage of cold storage of formaldehyde mixtures is that below 18 degrees Celsius formaldehyde will polymerize to paraformaldehyde. Also fixing at low temp. will dramatically increase fixation time (binding time) and might lead to less good morphology of the deeper lying tissue.
Kind regards, Dries
It depend on the nature of your sample and the recipe of the FAA that you used, because there are various recipes for FAA. If you sample is the type that become harden easily then reduce the concentration of the alcohol and formalin.
I used to use a FAA (Davidson's) for a limited range of animal tissue. It is a rapid fixative when compared to 10%NBF. Prolonged fixation hardened the tissue so we routinly changed the tissue into 70% aclcoho after 24 hours.l
The length of time your sample spend in the FAA depend on the nature of your sample, the size of the sample and the composition of the FAA. For example FAA can be composed as follows
(1) 50ml of 50% ethanol
25ml of 40% formalin
25ml of acetic acid
(2) 50ml of 70% ethanol
25ml of 40% formalin
25ml of acetic acid
(3) 70ml of 50% ethanol
15ml of 40% formalin
15ml of acetic acid
These are just examples of different composition of FAA ,there are lot of recipes for FAA. You will choose a composition that is suitable for your sample bearing in mind the nature of your sample. Do not forget that higher concentration of ethanol, large volume of formalin and acetic acid makes sample harden and become brittle.
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