5-HT (Serotonin) 2A Receptor Antibody

24288
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  • Refer to Dr. Schwendt's review
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Price: $400.00
Product ID : 24288

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Description

The antibody is provided as 100 uL of affinity purified serum in PBS (0.02 M sodium phosphate with 0.15 M sodium chloride, pH 7.5) with 1% BSA (bovine serum albumin), and 0.02% sodium azide.  Reacts with mouse, rat, and rabbit species.

The images above labeled "ImmunoStar Rabbit 5-HT 2A receptor antibody" are the results of staining of the mouse 5-HT 2A receptor in the brain, courtesy of Dr. Magdalena Zaniewska, Max-Delbruck-Centrum fur Molekulare Medizin Berlin, Germany.

The ImmunoStar 5HT 2A receptor antibody was quality control tested using standard immunohistochemical methods. The antiserum demonstrates strongly positive labeling of rat cortex, amygdala and hippocampus using indirect immunofluorescent and biotin/avidin-HRP techniques. Recommended primary dilutions are 1/300 - 1/500 in PBS/0.3% Triton X-100 - Bn/Av-HRP Technique . The addition of intensifying reagents such as nickel ammonium sulfate to the chromogen solution will approximately double the dilution factor as recommended.

Immunolabeling is completely abolished by preadsorption with synthetic rat 5HT2A receptor (22-41). Immunolabeling of Western blot revealed a single band of approximately 53kD.

Photo Description:  Low magnification IHC image of neurons staining for the 5-HT2A receptor in the rat cortex (top of page) and image of neuronal expression of the receptor in the amygdala (below). The bottom right photo is of the cortex.  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 1:500 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.

5HT2A Amygdala #24288   5HT2A Cortex, #24288
   

Host: Rabbit

Quantity / Volume: 100 uL

State: Liquid

Reacts With: Human, Marine, Mouse, Rabbit, Rat

Alternate Names:  5-hydroxytryptamine receptor 2A; 5Ht-2; 5-hydroxytryptamine (serotonin) receptor 2A, G protein-coupled, anti-5-HT 2A

Gene Symbol / ID, Accession #:  Htr2a,29595

RRID:  AB_572211

Reviews

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Reviewed by user
03/11/2014 - 10:36:22 AM
Only your AB shows a specific signal

(Please refer to the above western blot image submitted by Dr. Schwendt.)

We have been interested in detecting changes of 5-HT2a in rodent brain, primarily by immunoblotting, also by immunohistochemistry. We have tested several commercial antibodies, with only your Ab showing a specific signal, as verified by brain distribution, preadsorbtion and by a lack of signal in KO animals.

We have optimized a immunoblotting protocol which works well to detect 5-HT2a in rat brain lysates. Mouse tissue is a little bit more difficult but still possible

Immunoblotting: Rat or mouse brain samples were lysed by brief sonication in a buffer containing: 1% SDS in 1xPBS, protease inhibitors (Complete Mini, Roche). Samples were mixed with a modified Leamli buffer (containing a reducing agent; 40mM DTT), incubated at 37C for 30mins and separated by SDS-PAGE. Separated proteins were then blotted onto a PVDF membrane, blocked with 5% milk/TBS/0.1% Tween80 and probed with anti-5HT2a antibody (1:500, Immunostar #24288) followed by anti-rabbit HRP- conjugated secondary antibody (1:5000, Jackson Immuno). Signal was visualized using ECL2 kit according to manufacturer's instructions (Thermo).

Image Samples: 1 (Rat prefrontal cortex, 25ug), 2 (Rat laterodorsal thalamus, 25ug), 3 (Wt mouse cortex, 25 ug), 4 (5-HT2a KO mouse cortex, 25 ug), 5 (Wt mouse cortex, 75 ug), 6 (5-HT2a KO mouse cortex, 75 ug). As expected, sample in lane 2 showed low expression, and samples in lanes 4 and 6 showed no expression of 5-HT2a receptors. Mouse tissues obtained from Dr. Gonzalez-Maeso (Mount Sinai Hospital, NY) indicates detected 5Ht2a receptor signal.


Submitted by:

Marek Schwendt, PhD
Psychology Department
University of Florida
Gainesville, Fl



Reviewed by user
11/15/2012 - 02:37:02 PM
Antibodies are specific for mouse receptor

[The two pictures above labeled "ImmunoStar's Rabbit 5-HT 2A receptor antibody" are courtesy of Dr. Magdalena Zaniewska.]

Here are the results of staining of the mouse 5-HT 2A receptors in the brain with the use of Immunostar's antibodies (Cat no: 24288). I think the antibodies are specific for the mouse receptor (no receptor labeling in the brain of 5-HT 2A receptor knockout mouse) and can be used in a Western blot. What I already found out is that 5-HT 2A receptors in the cortex seem to be glycosylated (band around 75 kDa) since incubation with PNGaseF releases the protein that is around 50 kDa.

We received the 5-HT 2A receptor knockout animals from Dr. Joelle Adrien from another research team in Paris (see below).


Reviewed by:

Dr. Magdalena Zaniewska
Molekularbiologie von Hormonen im Herz-Kreislaufssystem
Max-Delbrϋck-Centrum fϋr Molekulare Medizin
Berlin, Germany

Dr. Joelle Adrien
Directeur de Recherches a l'Inserm
Faculte de Medecine Pierre et Marie Curie - Site Pitie Salpetriere
Paris, France

Publications

PROD # ANTIBODY CITATION SPECIES REACTIVITY APPL METHOD RESEARCH AREA # CITED BY PUBMED ID
24288 5-HT (Serotonin) 2A Receptor Antibody Fay, Richard, and Leszek Kubin. "5-HT 2A receptor-like protein is present in small neurons located in rat mesopontine cholinergic nuclei, but absent from cholinergic neurons." Neuroscience letters 314.1 (2001): 77-81. Rat 19 11698151
24288 5-HT (Serotonin) 2A Receptor Antibody Vis Neurosci. 1999 Mar-Apr;16(2):221-30. 5-HT2a receptors in the rabbit retina: potential presynaptic modulators. Pootanakit K1, Prior KJ, Hunter DD, Brunken WJ. Rabbit IHC 10367957
24288 5-HT (Serotonin) 2A Receptor Antibody Coulon P1, Kanyshkova T, Broicher T, Munsch T, Wettschureck N, Seidenbecher T, Meuth SG, Offermanns S, Pape HC, Budde T, Front Cell Neurosci. 2010 Oct 25;4:132. doi: 10.3389/ fncel.2010.00132. eCollection 2010. "Activity Modes in Thalamocortical Relay Neurons are Modulated by G(q)/G(11) Family G-proteins - Serotonergic and Glutamatergic Signaling" Mouse IF 21267426
24288 5-HT (Serotonin) 2A Receptor Antibody Kwan, Marilyn L., et al. "Airway vasculature after mycoplasma infection: chronic leakiness and selective hypersensitivity to substance P." American Journal of Physiology-Lung Cellular and Molecular Physiology 280.2 (2001): L286-L297. Rat IHC 24 11159008
24288 5-HT (Serotonin) 2A Receptor Antibody Vázquez DM1, Eskandari R, Zimmer CA, Levine S, López JF, Psychoneuroendocrinology. 2002 Jan-Feb;27(1-2):245-72. "Brain 5-HT receptor system in the stressed infant rat: implications for vulnerability to substance abuse" Rat ICC 56 11750782
24288 5-HT (Serotonin) 2A Receptor Antibody Itoh, Takeo, and Junko Kajikuri. "Characteristics of the actions by which 5?HT affects electrical and mechanical activities in rabbit jugular vein." British journal of pharmacology 164.3 (2011): 979-991. Rabbit IF 2 21449974
24288 5-HT (Serotonin) 2A Receptor Antibody Maekawa, Takashi, et al. "Characteristics of the actions by which 5?hydroxytryptamine affects electrical and mechanical activities in rabbit jugular vein graft." British journal of pharmacology 166.4 (2012): 1419-1432. Rabbit IF 4 22251164
24288 5-HT (Serotonin) 2A Receptor Antibody Rasul, Aram, et al. "Chronic mild stress modulates 5-HT1A and 5-HT2A receptor expression in the cerebellar cortex of NC/Nga atopic-like mice." Archives of dermatological research 305.5 (2013): 407-413. Mouse IHC 0 23440396
24288 5-HT (Serotonin) 2A Receptor Antibody J Vasc Surg. 2009 Sep;50(3):617-25. doi: 10.1016/j.jvs.2009.05.065. Chronic treatment of hydroxytryptamine type 2a receptor antagonist sarpogrelate hydrochloride modulates the vasoreactivity of serotonin in experimental rabbit vein grafts. Kodama A1, Komori K, Kajikuri J, Itoh T. Rabbit IF 19700096
24288 5-HT (Serotonin) 2A Receptor Antibody Volgin, Denys V., Georg M. Stettner, and Leszek Kubin. "Circadian dependence of receptors that mediate wake-related excitatory drive to hypoglossal motoneurons." Respiratory physiology & neurobiology 188.3 (2013): 301-307. Rat 0 23665050
24288 5-HT (Serotonin) 2A Receptor Antibody Pearlstein, E., et al. "Contribution of 5?HT to locomotion Mouse IHC 5 21501257
24288 5-HT (Serotonin) 2A Receptor Antibody Zeng, Fanqing, Chen Xu, and Ge Xu. "Distribution of serotonin 5-HT2A and 5-HT7 receptors in the Onuf's nucleus of the rat spinal cord." Neural Regen Res 3.8 (2008): 853-857. Rat 2 not found
24288 5-HT (Serotonin) 2A Receptor Antibody Rasul, Aram, et al. "Effect of chronic mild stress on serotonergic markers in the skin and brain of the NC/Nga atopic-like mouse strain." Archives of dermatological research 303.9 (2011): 625-633. Mouse 9 21400247
24288 5-HT (Serotonin) 2A Receptor Antibody Br J Pharmacol. 2010 Jul;160(5):1092-104. doi: 10.1111/j.1476-5381.2010.00753.x. Enhancement of mesenteric artery contraction to 5-HT depends on Rho kinase and Src kinase pathways in the ob/ob mouse model of type 2 diabetes. Matsumoto T1, Kobayashi T, Ishida K, Taguchi K, Kamata K. Mouse WB 20590603
24288 5-HT (Serotonin) 2A Receptor Antibody Exp Neurol. 2013 Sep;247:572-81. doi: 10.1016/j.expneurol.2013.02.008. Epub 2013 Feb 26.Grafting of fetal brainstem 5-HT neurons into the sublesional spinal cord of paraplegic rats restores coordinated hindlimb locomotion.Sławińska U1, Miazga K, Cabaj AM, Leszczyńska AN, Majczyński H, Nagy JI, Jordan LM. Rat 1 23481546
24288 5-HT (Serotonin) 2A Receptor Antibody Weber, Elaine T., and Rodrigo Andrade. "Htr2a gene and 5-HT2A receptor expression in the cerebral cortex studied using genetically modified mice." Frontiers in neuroscience 4 (2010). Mouse 31 20802802
24288 5-HT (Serotonin) 2A Receptor Antibody Hayashi, Takayuki, et al. "Intracerebroventricular fluvoxamine administration inhibited pain behavior but increased Fos expression in affective pain pathways." Pharmacology Biochemistry and Behavior 91.3 (2009): 441-446. IHC 6 18817806
24288 5-HT (Serotonin) 2A Receptor Antibody Stettner, Georg M., Leszek Kubin, and Denys V. Volgin. "Loss of motoneurons in the ventral compartment of the rat hypoglossal nucleus following early postnatal exposure to alcohol." Journal of chemical neuroanatomy 52 (2013): 87-94. Rat 0 23932955
24288 5-HT (Serotonin) 2A Receptor Antibody Fay, Richard, and Leszek Kubin. "Pontomedullary distribution of 5?HT2A receptor?like protein in the rat." Journal of Comparative Neurology 418.3 (2000): 323-345. Rat 96 10701830
24288 5-HT (Serotonin) 2A Receptor Antibody Bras H1, Gaytán SP, Portalier P, Zanella S, Pásaro R, Coulon P, Hilaire G, Eur J Neurosci. 2008 Sep;28(6):1097-107. doi:0.1111/j.1460- 9568.2008.06407.x. Epub 2008 Sep 9. "Prenatal activation of 5-HT2A receptor induces expression of 5-HT1B receptor in phrenic motoneurons and alters the organization of their premotor network in newborn mice". Mouse ICC 12 18783379
24288 5-HT (Serotonin) 2A Receptor Antibody Karaki, Samah, et al. "Quantitative Phosphoproteomics Unravels Biased Phosphorylation of Serotonin 5-HT2A Receptor at Ser280 by Hallucinogenic versus non-Hallucinogenic Agonists." Molecular & Cellular Proteomics (2014): mcp-M113. 0 not found
24288 5-HT (Serotonin) 2A Receptor Antibody Kong, Xiang-Yu, et al. "Robust upregulation of serotonin 2A receptors after chronic spinal transection of rats: An immunohistochemical study." Brain research 1320 (2010): 60-68. Rat IF 13 20085755
24288 5-HT (Serotonin) 2A Receptor Antibody Van Steenwinckel, Juliette, et al. "Role of spinal serotonin 5-HT2A receptor in 2?, 3?-dideoxycytidine-induced neuropathic pain in the rat and the mouse." Pain 137.1 (2008): 66-80. Rat, Mouse 27 17888573
24288 5-HT (Serotonin) 2A Receptor Antibody Rasul, Aram. "Serotonergic mechanisms in atopic dermatitis." (2012). 0 not found
24288 5-HT (Serotonin) 2A Receptor Antibody Xu, Chen, et al. "Serotonin 5?HT2A and 5?HT5A receptors are expressed by different motoneuron populations in rat Onuf's nucleus." Journal of Comparative Neurology 502.4 (2007): 620-634. Rat IHC 23 17394137
24288 5-HT (Serotonin) 2A Receptor Antibody Thibault, Karine, et al. "Serotonin 5-HT2A receptor involvement and Fos expression at the spinal level in vincristine-induced neuropathy in the rat." Pain 140.2 (2008): 305-322. Rat 32 18930597
24288 5-HT (Serotonin) 2A Receptor Antibody Loyd, Dayna R., et al. "Sex differences in serotonin enhancement of capsaicin-evoked calcitonin gene-related peptide release from human dental pulp." Pain 153.10 (2012): 2061-2067. Human 6 22819536
24288 5-HT (Serotonin) 2A Receptor Antibody Neuropsychopharmacology. 2009 Jan;34(2):410-23. doi: 10.1038/npp.2008.71. Epub 2008 Jun 4. Stress impairs 5-HT2A receptor-mediated serotonergic facilitation of GABA release in juvenile rat basolateral amygdala. Jiang X1, Xing G, Yang C, Verma A, Zhang L, Li H. Rat IF 18536707
24288 5-HT (Serotonin) 2A Receptor Antibody Dumitrascu, R., et al. "Terguride ameliorates monocrotaline-induced pulmonary hypertension in rats." European Respiratory Journal 37.5 (2011): 1104-1118. Rat IHC 44 20947677
24288 5-HT (Serotonin) 2A Receptor Antibody Van Steenwinckel, J., et al. "The 5-HT2A receptor is mainly expressed in nociceptive sensory neurons in rat lumbar dorsal root ganglia." Neuroscience 161.3 (2009): 838-846. Rat IHC 27 19362128
24288 5-HT (Serotonin) 2A Receptor Antibody J Comp Neurol. 2004 May 10;472(4):496-511. The 5-HT2A receptor is widely distributed in the rat spinal cord and mainly localized at the plasma membrane of postsynaptic neurons. Doly S1, Madeira A, Fischer J, Brisorgueil MJ, Daval G, Bernard R, Vergé D, Conrath M. Rat ICC 68 15065122
24288 5-HT (Serotonin) 2A Receptor Antibody Neurochem Res. 2005 un-Jul;30(6-7):825-37. The pre-nervous serotonergic system of developing sea urchin embryos and larvae: pharmacologic and immunocytochemical evidence.Buznikov GA1, Peterson RE, Nikitina LA, Bezuglov VV, Lauder JM. Marine IHC 16187217
24288 5-HT (Serotonin) 2A Receptor Antibody Kong, X-Y., et al. "The time course of serotonin 2A receptor expression after spinal transection of rats: an immunohistochemical study." Neuroscience 177 (2011): 114-126. Rat 9 21211552

Q & A Wall

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I would be very grateful if you could send me the Western Blot protocol for 5-HT2A R Ab. Did you prepare the WB from the brain tissue homogenate?
Question By Admin

Western blots of 5HTRs is challenging. We have examined crude homogenates and immunopurified tissue extracts. The extraction buffer was RIPA buffer with enzyme inhibitor tablets added (Sigma). We used the Pierce Seize X mammalian immunoprecipitation kit 45210.


Due to the difficulty in running westerns with this antibody, ImmunoStar does not support or warrant the western blot application. The western information is included as specificity information only.

For those that wish to proceed with the western blot, here is a summary of our protocol for the 5-HT2A antibody:

1. Tissue is homogenized in either electrophoresis sample bufferor (in which case you go straight to electrophoresis with no cleanup) or in 0.05 Mis-saline with 10% sucrose and 1 mM PMSF, pH 7.6 (in which case you proceed as below).
2. Centrifuge homogenized sample and collect pellet by centrifugation. Solubilize pellet in 20 mM tris-saline, pH 8, 1 mM EDTA, and 1% deoxycholate, sonicate and incubate hr on ice. Centrifuge and collect supernatant. Subject supernatant to electrophoresis.
3. Electrophorese and transfer onto nitrocellulose by standard methods.
4. Block membrane in blotto buffer (TBS with 0.05% Tween 20 and 0.2% I-block [Tropix #AI300]. This is also the antibody incubation buffer.
5. Incubate blots in rab anti 5HT2a at 1:500 overnight in refrigerator.
6. Proceed with chemiluminescent detection (Amersham ECL kit)


Answer By Admin on 2015-06-11 04:37:52

We would like to know if this antibody cross-reacts with human. We want to use it on sections for immunohistochemistry.
Question By Admin

Our antibody was raised against rat 22-41. This sequence blasted and got 78% similarity to human, though not a lot of amino acids in a row to say it will definitely x-react. We don't know if anyone has used in human and published either.

Here's the alignment:

Rat GDPRLYHNDFNSRDANTSE

Hum DDTRLYSNDFNSGEANTSD

Answer By Admin on 2015-06-09 06:20:52

MSDS

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.

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