The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use at an assay dependent concentration. PubMed: 22474435
Use a concentration of 1 - 5 µg/ml. (see Ni et al). Fix with ice cold acetone. Dilute antibody in 1.5% blocking serum.
Use at an assay dependent concentration.
1/100 - 1/1000. Used on gelatin embedded perfusion fixed tissue (see recommended protocol).
Serotonin (5-hydroxytryptamine, or 5-HT) is a monoamine neurotransmitter synthesised in serotonergic neurons in the central nervous system and enterochromaffin cells in the gastrointestinal tract. Serotonin plays an important part in the biochemistry of depression, migraine, bipolar disorder and anxiety. It is also believed to be influential on sexuality and appetite. 5-HT is generally thought not to be released from synaptic terminal buttons in the manner of classical neurotransmission but from serotonergic varacosities into the extra neuronal space. From here it is free to diffuse over a relatively large region of space (>20µm) and activate 5-HT receptors located on the dendrites, cell bodies and presynaptic terminals of adjacent neurons. Serotonergic action is terminated primarily via uptake of 5-HT from the synapse. This is through the specific monoamine transporter for 5-HT, 5-HT reuptake transporter, on the presynaptic neuron. The pharmacology of 5-HT is extremely complex, with its actions being mediated by a large and diverse range of 5-HT receptors.
ab6336 in rat brain tissue fixed with 4% paraformaldehyde. The staining is appropriate, lighting up cell bodies, dendrites and terminals of known 5-HT containing neurones. These pictures are from the review provided by Jim Deuchars.
IHC image of Serotonin staining in mouse brain (coronal) formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab6336, 5µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
Immunohistochemistry (Frozen sections) - Anti-Serotonin antibody [YC5/45] (ab6336)This image is courtesy of Ms. Szu-yu Liu by an abreview
Paraformaldehyde-fixed rabbit spinal cord tissue stained for Serotonin using ab6336 at 1/200 dilution in immunohistochemical analysis. Power Block™ Universal Blocking Reagen t(Biogenex) was used as ablocking agent for 10 minute(s) at a concentration of 10% at 37°C. Biotin-SP-conjugated Goat Anti-Rat IgG was used as the secondary antibody at 1/500 dilution.
Sitaraman D et al. Discrete Serotonin Systems Mediate Memory Enhancement and Escape Latencies after Unpredicted Aversive Experience inDrosophilaPlace Memory. Front Syst Neurosci11:92 (2017).
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Dhillon RS et al. Axonal plasticity underpins the functional recovery following surgical decompression in a rat model of cervical spondylotic myelopathy. Acta Neuropathol Commun4:89 (2016).
Read more (PubMed: 27552807) »