Fixation of transsynaptically transported WGA-HRP and fluorescent dyes used in combination.
The transsynaptic transport of WGA-HRP is a powerful tool for the identification of last order interneurones in the spinal cord, but differentiation of primarily labelled motoneurones from transsynaptically labelled interneurones can be difficult. We therefore combined the transsynaptic labelling of interneurones with WGA-HRP and the labelling of motoneurones with fluorescent dyes (fluorogold, red beads, fast blue, DAPI) by injecting a mixture of the tracers into the nervi mediani of rats. The influence of different fixatives on the simultaneous preservation of WGA-HRP and fluorescence is evaluated. It is shown that, of several perfusion protocols, only perfusion with paraformaldehyde in addition to 1.4% lysine and 0.23% periodate allowed simultaneous visualisation of transsynaptic WGA-HRP and fluorescent dyes.
Herzog
J
J
Department of Physiology, Christian-Albrechts-Universität zu Kiel, Olshausenstrasse 40, D-24098, Kiel, Germany.
Kümmel
H
H
D000079:Acetaldehyde / Q000493:pharmacokinetics; D000171:Acrolein / Q000494:pharmacology; D000818:Animals; D001370:Axonal Transport / Q000187:drug effects* / Q000502:physiology; D001881:Borates / Q000494:pharmacology; D002021:Buffers; D005404:Fixatives / Q000494:pharmacology; D005456:Fluorescent Dyes / Q000378:metabolism; D005557:Formaldehyde / Q000494:pharmacology; D005976:Glutaral / Q000494:pharmacology; D007395:Interneurons / Q000166:cytology / Q000378:metabolism; D008239:Lysine / Q000494:pharmacology; D008475:Median Nerve / Q000166:cytology / Q000378:metabolism / Q000601:surgery; D009046:Motor Neurons / Q000166:cytology / Q000378:metabolism; D009415:Nerve Net / Q000166:cytology* / Q000378:metabolism; D009434:Neural Pathways / Q000166:cytology* / Q000378:metabolism; D010504:Periodic Acid / Q000494:pharmacology; D011108:Polymers / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D013116:Spinal Cord / Q000166:cytology / Q000378:metabolism; D009435:Synaptic Transmission / Q000187:drug effects* / Q000502:physiology; D016707:Tissue Fixation / Q000379:methods* / Q000592:standards; D018998:Wheat Germ Agglutinin-Horseradish Peroxidase Conjugate / Q000378:metabolism
D001881:Borates; D002021:Buffers; D005404:Fixatives; D005456:Fluorescent Dyes; D011108:Polymers; D018998:Wheat Germ Agglutinin-Horseradish Peroxidase Conjugate; C046311:periodate-lysine-paraformaldehyde; D010504:Periodic Acid; D005557:Formaldehyde; D000171:Acrolein; C010634:sodium borate; D000079:Acetaldehyde; D008239:Lysine; D005976:Glutaral; C003043:paraform