Hi Yuxiang,TH double labeling definitely works with many different markers. The key will be to have a TH antibody and a dendrite/axon marker that are raised in different species. This should be no major obstacle. There are several excellent TH antibodies raised in different hosts that are available.  The same is true for MAP and Tau antibodies. (I'm not sure if non-mouse acetylated alpha tubulin antibodies are out there. The antibody I'm familiar with is a mouse monoclonal). The best news is that these antibodies and antigens tend to yield robust staining so you should be able to assess double labeling accurately.

dave

To my knowledge nigrostriatal dopaminergic neurons are immunostained by TH antibody; it labels the soma, the axons and dendrites

Thank you Dr. Zaccone!

So, is there anyway to tell if what I stained are axons or dendrites? I can't see any spine morphology in these TH positive neuritis.

There are several markers that could allow you to distinguish axons and dendrities. Acetelyated alpha tubulin and Tau are classic axonal markers and MAP2 is a classic marker for dendrites.

Thank you David. While these markers can generally distinguish axons and dendrites, the question is I want to specifically tell if the TH positive "dopaminergic neurite" is an axon or a dendrite.

Not sure if double staining works as there are tons of neurites in the area. Will try. Thanks!

Dear Yuxiang,

I also located axon hillocks using AcT in the intracardiac neurons of the cod, lying in the pacemaker(unpublished).See Aransay et al 2015 paper Frontiers in Neuroanatomy-VTA cells.They  used TH immunostaining to locate cell soma,axons and dendrites,but used different neuronal markers.

Giacomo

Hi Yuxiang,TH double labeling definitely works with many different markers. The key will be to have a TH antibody and a dendrite/axon marker that are raised in different species. This should be no major obstacle. There are several excellent TH antibodies raised in different hosts that are available.  The same is true for MAP and Tau antibodies. (I'm not sure if non-mouse acetylated alpha tubulin antibodies are out there. The antibody I'm familiar with is a mouse monoclonal). The best news is that these antibodies and antigens tend to yield robust staining so you should be able to assess double labeling accurately.

dave

Hi Yuxiang, neurofilament protein providing structural support of axons as well as calbindin DK 28 should be also reliable markers to use antibody pairs with TH antibodies.In particular  calbindin DK 28 is a marker for cell bodies, dendrites, spines and matrix of cytoplasm.As fish neurobiologist I found that intrinsic cardiac neurons (ICN) in fish are immunolabeled with TH antibodies although these neurons are unipolar, but they synapse with TH positive preganglionic nerve fibers.

Giacomo