Medizinische Universität Graz Austria/Österreich - Forschungsportal - Medical University of Graz
Gewählte Publikation:
SHR Neuro Krebs Kardio Lipid Stoffw Microb
Di Biase, V; Flucher, BE; Obermair, GJ.
Resolving sub-synaptic compartments with double immunofluorescence labeling in hippocampal neurons.
J Neurosci Methods. 2009; 176(2):78-84
Doi: 10.1016/j.jneumeth.2008.08.025
Web of Science
PubMed
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- Führende Autor*innen der Med Uni Graz
- Di Biase Valentina
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- Abstract:
- Immunofluorescence microscopy of synaptic proteins is a powerful and commonly used approach in cellular neurosciences. Many studies use green/red color overlays of immunofluorescence images to demonstrate synaptic co-localization of an unknown protein with a known synaptic marker. However, this approach fails to identify the specific sub-synaptic compartment in which a protein is localized. Here we describe a novel analysis method to determine the precise location of proteins within synapses of cultured hippocampal neurons with double immunofluorescence staining. This approach is based on center-to-center distance measurements of fluorescent clusters of protein pairs coexisting in synapses. We validated the method by analyzing the distances between different combinations of well-established synaptic marker proteins. The results demonstrate that protein pairs in the active zone and the postsynaptic density, two sub-synaptic compartments which are separated by less than 50 nm, can be readily distinguished from each other and from marker pairs co-localized within a single sub-synaptic compartment. Thus, center-to-center distance analysis can resolve the distance across the synaptic cleft and it is useful for localizing synaptic proteins to specific pre- and postsynaptic compartments.
- Find related publications in this database (using NLM MeSH Indexing)
- Animals -
- Cells, Cultured -
- Embryo, Mammalian -
- Fluorescent Antibody Technique - methods
- Hippocampus - cytology
- Image Processing, Computer-Assisted -
- Mice -
- Mice, Inbred BALB C -
- Microscopy, Fluorescence - methods
- Nerve Tissue Proteins - metabolism
- Neurons - cytology
- Presynaptic Terminals - chemistry Presynaptic Terminals - metabolism
- Synapses - chemistry Synapses - metabolism Synapses - physiology
- Find related publications in this database (Keywords)
- Immunocytochemistry
- Presynaptic
- Subcellular localization
- Microscopy
- Piccolo
- RIM
- Synapsin
- PSD-95
- vGLUT1