I was thinking of using ethanol/methanol. Does anyone have a whole animal alcohol perfusion procedure? Is this possible?
You're missing the whole point of fixation here. Fixation aims to stop autolysis and preserve cells/tissues in as original a state as possible. This is mostly by affecting the proteins: alcohols will coagulate proteins, aldehydes stabilize and then later crosslink, etc. If you fix the cells, you fix the ECM.
The only way I know to recover cells from organs is to digest the ECM (e.g. with collagenase) and running them through a cell strainer. This will not net you a quantitative amount of cells however! Plenty of cells will be destroyed in the process.
I think you're better off figuring out a method of weighing/measuring the organ accurately, then estimating average cell size on histologic sections and working out a ratio that way to determine the amount of cells.
Btw, don't use alcohols to fix tissue for histology, the morphology will be horrible. Formaldehyde-based fixatives are generally the better option for decent results (but not for your health, so use caution).
Hi Michael, I think that fixation is not the best way to reach your goal. I agree with Jonathan, if you want to count cells of your organ you have to digest it with collagenase, but you will destroy many cells!
The best way to have a quantitative value is to measure the weigh first, then fix with PFA 4% for 12- 24 hours, then dehydrate and include in paraffine and cut sections to count in each section cells making an average of the total number X the number of sections.
Good luck!
Lorena
For whole organ you can use Carnoy's or Bouin's fixative.Only Bouin's fixative is not so good for fixation of kidney.Good results are received when fixation is carried out in low temperature(o or +6).It's connected with stoping autolitic process.
- Badges
- Science topic
- Similar topics
- Biological Science
- Histology