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The ImmunoStar serotonin (5HT) transporter was quality control tested using standard immunohistochemical methods. The antiserum demonstrates strongly positive labeling of rat raphe nuclei, hypothalamus, cortex and spinal cord using indirect immunofluorescent and biotin/avidin-HRP techniques. Recommended primary dilution is 1/10,000 - 1/15,000 in PBS/0.3% Triton X-100 - Biotin/avidin-HRP.
By Western blot analysis using rat brain extracts of cortex, hypothalamus, midbrain, and hindbrain, the antibody specifically labels a single band. The predicted molecular weight of serotonin transporter based on its nucleic acid sequence is 67,815. The actual weight that one might observe from tissue extracts could be higher due to glycosylation. We validated the antibody with rat SERT overexpressed in an insect cell expression system. The MW from that was about 67 kD. Our SERT antibody dilution was 1:20,000 using chemiluminescent detection.
Immunolabeling is completely abolished by pre-adsorption with synthetic rat 5HT transporter (602-622).
Photo Description: IHC image of neurons staining for the 5-HT transporter in the raphe nucleus of the rat brainstem (above). The tissue was fixed with 4% formaldehyde in 0.1 M phosphate buffer, before being removed and prepared for vibratome sectioning. Floating sections were incubated at RT in 10% goat serum in PBS, before standard IHC procedure. Primary antibody was incubated at 1:10000 for 48 hours, goat anti-rabbit secondary was subsequently added for 1 hour after washing with PBS. Light microscopy staining was achieved with standard biotin-streptavidin/HRP procedure and DAB chromogen. Florescent secondary (below)
Quantity / Volume: 100 µL
State: Lyophilized Whole Serum
Availability: In Stock
Reacts With: Human, Marine, Monkey, Mouse, Rat, Sparrow
Alternate Names: 5HTT; HTT; Sodium-dependent serotonin transporter; Solute carrier family 6 member 4; Slc6a4; solute carrier family 6 (neurotransmitter transporter, serotonin), member 4
Immunogen: Rat 5-HT transporter (579-599)
Gene Symbol / ID, Accession #: Slc6a4,25553
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Q & A Wall
First, is the antibody specific to SERT, or is there affinity for DAT or NET? Also, can the antibody be used for mouse and human as well as for rat? Has this antibody been used for immunoprecipitation?
Question By Michael W
The antibody is specific for SerT and does not cross react with DAT or NET.
The antiserum was characterized anatomically by Western blot and by preadsorption with a SerT specific peptide. A BLAST search of the peptide sequence finds only SerT from different species. The sequence is 100% homologous with rat (because that's what was chosen to raise it against).
It is 90% homologous with mouse so it will probably work. It is 85% homologous with human so it might work, but we haven't tried it.
The antibody has not been used for immunoprecipitation; not that it wouldn't work, but we've not tried it. We have used Vector ABC kits with nice results and secondary fluorescent antisera such as Cy3, also with nice results. Thinner sections work best since there are numerous fine fibers in many areas of the CNS.
Answer By Tech Support on 2006-09-22 13:44:20
The product specification sheet suggests a single band reactivity of SERT to multiple regions of rat brain on western blots. I am wondering if you could perhaps please share the western blot images showing the performance of this antibody to brain extracts of rat as mentioned in the spec sheet. We were unable to find any western blot images in the references listed on the website.
Question By Chandana
I looked through our library of references but unfortunately could not find an article in which rat tissue was used with western blot using the 5-HT Transporter antibody. However I have information from our lab from a few years ago:
The predicted molecular weight of serotonin transporter based on its nucleic acid sequence is 67,815. The actual weight that one might observe from tissue extracts could be higher due to glycosylation. We validated the antibody with rat SERT overexpressed in an insect cell expression system. The MW from that was about 67 kD. Our SERT antibody dilution was 1:20,000 using chemiluminescent detection.
Answer By Colleen on 0000-00-00 00:00:00
This product contains the preservative sodium azide. The concentration percent of the sodium azide is ≤ .09%. Although this hazardous substance is a concentration below that required for the preparation of a Material Safety Data Sheet, we created a standard MSDS for your records.Download MSDS