Anti-mouse 5-HT_1A receptor, rat monoclonal antibody (4A6)
Anti-mouse 5-HT_2C receptor, rat monoclonal antibody (6D2)

5-HT_1Areceptor and 5-HT_2C receptor are G protein-coupled receptors activated by serotonin (5-HT). Both are localized mainly in the central nervous system, and their functions controlling memory, food intake, sleep, pleasure, and anxiety, etc. Anxiolytics and antipsychotics acting on these receptors have been developed, and they are attracting attention of researchers as novel targets for drug discovery.

This product is a rat monoclonal antibody established by DNA immunization and specifically recognizes native forms of 5-HT_1A receptor and 5-HT_2C receptor.

Features

• Compatible with immunohistochemical staining
• Specifically recognizes native forms of 5-HT_1Areceptor or 5-HT_2C receptor
• Rat monoclonal antibodies established by DNA immunization

特征

• 与免疫组化染色兼容
• 特异性识别天然形式的 5-HT _1A受体或 5-HT _2C受体
• DNA免疫法建立的大鼠单克隆抗体

Description

DNA immunization

What is DNA immunization?

DNA immunization is a technique for generating antibodies against target proteins by using antigen proteins expressed in the animal body with expression vectors containing the target genes.

Features of DNA immunization

• Yields Antibodies specifically recognizing native forms of proteins
• Suitable for preparation of antibodies recognizing membrane proteins that were difficult to prepare by conventional methods

DNA免疫的特点

• 产生特异性识别天然蛋白质形式的抗体
• 适用于制备识别常规方法难以制备的膜蛋白的抗体

Principles of DNA immunization

Usage example – Immunohistochemical staining

5-HT_1Areceptor antibody

• 

• 

• Localization in the cell body of prefrontal neurons was confirmed.
  This localization is consistent with the site highly expressing mRNA for 5-HT_1A.

5-HT_2C receptor

• 

• 

• Localization in the cell body of prefrontal neurons was confirmed.
  This localization is consistent with the site highly expressing mRNA for 5-HT_2C.

Experimental conditions

Specimen: Prefrontal area of the brain from 10-week-old wild-type mice
Sections: 12 µm-thick frozen sections
Activation condition: Microwave treatment in a citrate buffer (pH 7.0) for 10 minutes
Antibody concentration: 1 µg/mL

实验条件

标本：来自 10 周龄野生型小鼠的大脑前额叶区域
切片：12 µm 厚的冰冻切片
激活条件：在柠檬酸盐缓冲液 (pH 7.0) 中微波处理 10 分钟
抗体浓度：1 µg/mL

Usage example – Flow cytometry

5-HT_1A receptor antibody

• 

• An obvious shift was observed only for cells expressing 5-HT_1A.

Anti 5-HT_2C receptor antibody

• 

• 

• A clearer shift was observed for cells expressing 5-HT_2C, compared with the competitor.

Usage example – Immunohistochemical staining of the site expressing mRNA for 5-HT_1A

Positive signals were noted in amygdala (BLA), entorhinal cortex (Ent), interpeduncular nucleus (IP), and dorsal raphe nuclei (DRN).
This localization is consistent with the site highly expressing mRNA for 5-HT_1A.

Experimental conditions

Specimens: Various areas of the brain from 10-week-old wild-type mice
Sections: 12 µm-thick frozen sections
Activation condition: Microwave treatment in a citrate buffer (pH 7.0) for 10 minutes
Antibody concentration: 1 µg/mL

Usage example – Immunohistochemical staining of the site expressing mRNA for 5-HT_2C receptor

Positive signals were noted in nucleus accumbens shell (NAc shell), amygdala (BLA), and dorsal raphe nuclei (DRN).
This localization is consistent with the site highly expressing mRNA for 5-HT_2C.

Experimental conditions

Specimens: Various areas of the brain from 10-week-old wild-type mice
Sections: 12 µm-thick frozen sections
Activation condition: Microwave treatment in a citrate buffer (pH 7.0) for 10 minutes
Antibody concentration: 1 µg/mL

All immunohistochemical staining image data presented above are courtesy of Dr. Matsuda, Graduate School of Pharmaceutical Sciences, Osaka University in Japan and Dr. Takuma and Hasebe of Graduate School of Dentistry, Osaka University in Japan.

实验条件

标本：10 周龄野生型小鼠大脑的各个区域
切片：12 µm 厚的冰冻切片
激活条件：在柠檬酸盐缓冲液 (pH 7.0) 中微波处理 10 分钟
抗体浓度：1 µg/mL

以上所有免疫组化染色图像数据均由日本大阪大学药学研究生院 Matsuda 博士和日本大阪大学牙科研究生院的 Takuma 和 Hasebe 博士提供。

Anti-mouse serotonin transporter, rat monoclonal antibody (R5-3-2)

Serotonin transporter is a 12-transmembrane protein. It incorporates extracellular serotonin in the brain into the presynapse to modulate serotonin levels. With its reported involvement in sleep, fear, and anxiety, serotonin transporter has been investigated as a target of antidepressants.

Features

• Compatible with immunohistochemical staining
• Established by DNA immunization

Description

Usage example – Immunohistochemical staining (fluorescence staining)

Immunohistochemical staining was performed for specimens from dorsal raphe nuclei (DRN) rich in serotonin neurons where expression of serotonin transporter has been reported. As a result, fluorescence signals were noted in the neuronal cell body.

Experimental conditions

• Samples: Frozen sections of the dorsal raphe nuclei from the brain of 10-week-old ICR mice
• Staining method: Fluorescent antibody staining
• Activation condition: Microwave treatment in a citrate buffer (pH 6.0) for 10 minutes
• Antibody concentration: 1:100
• Primary antibody: This product
• Secondary antibody: Alexa 594 goat anti Rat IgG

Image data are courtesy of Dr. Takuma and Hasebe of Graduate School of Dentistry, Osaka University.

Protocol example

1. Wash the tissue section on the slide glass with phosphate-buffered saline containing 0.3 % Triton X-100^® (PBST).
2. Perform activation treatment in a citrate buffer (pH 6.0) with a microwave oven (10 minutes).
3. Bring the tissue section back to ambient temperature.
4. Wash the tissue section with 0.3 % PBST.
5. Block the tissue section with 5% goat serum-PBS (1 hour).
6. Incubate the tissue section with the primary antibody (this product) diluted 100-fold with 0.3 %PBST (4°C, overnight).
7. Wash the tissue section with 0.3 % PBST.
8. Incubate the tissue section with the secondary antibody (room temperature, 2 hours).
9. After washing with 0.3% PBST, mount the tissue section.

Triton X-100 is a registered trademark of Dow Chemical Company.

Usage example – Immunohistochemical staining (DAB staining)

Fluorescence signals were noted in the neuronal cell body.

Experimental conditions Samples: Frozen sections of the dorsal raphe nuclei from the brain of 10-week-old ICR mice Staining method: Enzyme-labeled antibody method (DAB staining) Activation condition: Microwave treatment in a citrate buffer (pH 6.0) for 10 minutes Antibody concentration: 1:100

Image data are courtesy of Dr. Takuma and Hasebe, Graduate School of Dentistry, Osaka University in Japan.

Protocol example

1. Wash the tissue section on the slide glass with phosphate-buffered saline containing 0.3 % Triton X-100^® (PBST).
2. Perform activation treatment in a citrate buffer (pH 6.0) with a microwave oven (10 minutes).
3. Bring the tissue section back to ambient temperature.
4. Wash the tissue section with 0.3 % PBST.
5. Treat the tissue section with 40% methanol containing 0.3 % H₂O₂ (30 minutes)..
6. Block the tissue section with 5% goat serum-PBS (1 hour).
7. Incubate the tissue section with the primary antibody (this product) diluted 100-fold with 0.3 %PBST (4°C, overnight).
8. Wash the tissue section with 0.3 % PBST.
9. Incubate the tissue section with the biotinylated secondary antibody (room temperature, 2 hours).
10. Wash the tissue section with 0.3 % PBST.
11. Incubate the tissue section with DAB. After dehydration treatment, mount the tissue section.

Triton X-100 is a registered trademark of Dow Chemical Company.

Usage example – Flow cytometry

Green: 293T cells
Red: 293T cells expressing mouse serotonin transporter

Antibody concentration: 0.1 µg/mL

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