Perfusion with Tyrode's solution for CNS IHC
I have been attempting some rat brain IHC, and all methods in journal
articles I have read refer to perfusing with Ca-free Tyrode's solution
(a paraformaldehyde fixative of some description). This involves
anaesthetising the animal (usually with a barbiturate), cutting the
animal open and pumping fixative into a coronary vessel and draining
blood, while the animal is still alive (it's not alive for long). The
brain is then removed and placed in 4% paraformaldehyde to 'post-
fix'. I imagine the reason for perfusing is to get fixative to the
brain as quickly as possible, to minimise changes after death.
This method seems quite time-consuming, labour-intensive and
potentially distressing to the investigator or animal, not to mention
the introduction of a CNS-active drug which may affect the results.
Many parts of the brain are physically distant from blood supply.
Consequently I have been pondering the value of this type of
perfusion. Can any Histonetters comment on this? I have attempted
some preliminary experiments without perfusing (just post-fixing), and
labelling appears good. Would this deviation from the standard method
jeopardise the work's chances at publication?
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