Ms X Glutamic Acid Decarboxylase

Species Reactivity	Key Applications	Host	Format	Antibody Type
H, R	IHC, WB	M	Purified	Monoclonal Antibody

Species Reactivity

Key Applications

Host

Format

Antibody Type

H, R

IHC, WB

Purified

Monoclonal Antibody

Description
Catalogue Number	MAB351R
Brand Family	Chemicon®
Trade Name	Chemicon
Description	Anti-Glutamate Decarboxylase Antibody, 65 kDa isoform, clone GAD-6
Alternate Names	GAD65
Background Information	Gutamic acid decarboxylase (GAD; E.C. 4.1.1.15) is the enzyme responsible for the conversion of glutamic acid to gamma-aminobutyric acid (GABA), the major inhibitory transmitter in higher brain regions, and putative paracrine hormone in pancreatic islets. Two molecular forms of GAD (65kDa and 67kDa, 64% aa identity between forms) are highly conserved and both forms are expressed in the CNS, pancreatic islet cells, testis, oviduct and ovary. The isoforms are regionally distributed cytoplasmically in the brains of rats and mice (Sheikh, S. et al. 1999). GAD65 is an ampiphilic, membrane-anchored protein (585aa), encoded on human chromosome 10, and is responsible for vesicular GABA production. GAD67 is cytoplasmic (594aa.), encoded on chromosome 2, and seems to be responsible for significant cytoplasmic GABA production. GAD expression changes during neural development in rat spinal cord. GAD65 is expressed transiently in commissural axons around E13 but is down regulated the next day while GAD67 expression increases mostly in the somata of those neurons (Phelps, P. et al. 1999). In mature rat pancreas, GAD65 and GAD67 appear to be differentially localized, GAD65 primarily in insulin-containing beta cells and GAD67 in glucagon-containing (A) cells (Li, L. et al. 1995). GAD67 expression seems to be particularly plastic and can change in response to experimental manipulation (for example neuronal stimulation or transection) or disease progression and emergent disorders like schizophrenia (Volk et al., 2000). Colocalization of the two GAD isoforms also shows changes in GAD65/GAD67 distributions correlated with certain disease states such as IDDM and SMS.

Description

Catalogue Number

MAB351R

Brand Family

Chemicon®

Trade Name

Chemicon

Description

Anti-Glutamate Decarboxylase Antibody, 65 kDa isoform, clone GAD-6

Alternate Names

GAD65

Background Information

Gutamic acid decarboxylase (GAD; E.C. 4.1.1.15) is the enzyme responsible for the conversion of glutamic acid to gamma-aminobutyric acid (GABA), the major inhibitory transmitter in higher brain regions, and putative paracrine hormone in pancreatic islets. Two molecular forms of GAD (65kDa and 67kDa, 64% aa identity between forms) are highly conserved and both forms are expressed in the CNS, pancreatic islet cells, testis, oviduct and ovary. The isoforms are regionally distributed cytoplasmically in the brains of rats and mice (Sheikh, S. et al. 1999). GAD65 is an ampiphilic, membrane-anchored protein (585aa), encoded on human chromosome 10, and is responsible for vesicular GABA production. GAD67 is cytoplasmic (594aa.), encoded on chromosome 2, and seems to be responsible for significant cytoplasmic GABA production. GAD expression changes during neural development in rat spinal cord. GAD65 is expressed transiently in commissural axons around E13 but is down regulated the next day while GAD67 expression increases mostly in the somata of those neurons (Phelps, P. et al. 1999). In mature rat pancreas, GAD65 and GAD67 appear to be differentially localized, GAD65 primarily in insulin-containing beta cells and GAD67 in glucagon-containing (A) cells (Li, L. et al. 1995). GAD67 expression seems to be particularly plastic and can change in response to experimental manipulation (for example neuronal stimulation or transection) or disease progression and emergent disorders like schizophrenia (Volk et al., 2000). Colocalization of the two GAD isoforms also shows changes in GAD65/GAD67 distributions correlated with certain disease states such as IDDM and SMS.

Product Information
Format	Purified
Control	Brain tissue
Presentation	Lyophilized. Dissolve contents of vial in 100 µL of sterile, distilled water. This results in a final antibody concentration of 1 mg/ml in 10 mM potassium phosphate, 70 mM sodium chloride, pH 7.4 containing no preservatives.

Product Information

Format

Purified

Control

Brain tissue

Presentation

Lyophilized. Dissolve contents of vial in 100 µL of sterile, distilled water. This results in a final antibody concentration of 1 mg/ml in 10 mM potassium phosphate, 70 mM sodium chloride, pH 7.4 containing no preservatives.

Applications
Application	Anti-Glutamate Decarboxylase Antibody, 65 kDa isoform, clone GAD-6 is an antibody against Glutamate Decarboxylase for use in IH & WB.
Key Applications	Immunohistochemistry Western Blotting
Application Notes	Immunohistochemistry: (<= 1 μg/ml) Optimal working dilutions must be determined by end user. Immunohystochemical Staining Procedures The following procedure was developed to localize GAD in rat brain sections of cerebellum. Perform all steps at room temperature unless otherwise indicated. Where normal serum is indicated, use normal serum from the same species as the source of the secondary antibody.This procedure represents suggested guidelines for the use of anti-GAD. Fixation regimen, antibody concentrations, and incubation conditions for a given experimental system should be determined empirically. 1. Perfuse rats with 100 mM phosphate buffer, pH 7.4, containing 1% paraformaldehyde, 0.34% L-lysine, and 0.05% sodium m-periodate (1% PLP). 2. Postfix brains in 1% PLP for 1-2 hours. Longer fixation times may reduce labeling intensity. 3. Transfer brains to 100 mM phosphate buffer containing 30% sucrose, and gently agitate on a shaker platform at +4°C for 48-60 hours. 4. Using a sliding microtome, cut 30 mm sections of frozen cerebellum. As the sections are cut, collect them in a vial of cold 100 mM phosphate buffer. 5. Incubate sections in phosphate-buffered saline (PBS) containing 1.5% normal serum and 0.2% TritonX-100 for 30 minutes. 6. On a shaker platform, incubate sections with anti-GAD (diluted in PBS containing 1.5% normal serum and 0.2% Triton X-100 to a final antibody concentration of 1 mg/ml) for 12-36 hours at +4°C. 7. On a shaker platform, rinse sections eight times, 10-15 minutes per rinse, in PBS. 8. Detect with a standard secondary antibody detection system (Hsu et al., 1981; Falini & Taylor, 1983; Harlow & Lane, 1988; Taylor, 1978). 9. Mount sections, dehydrate, and apply coverslips.

Applications

Application

Anti-Glutamate Decarboxylase Antibody, 65 kDa isoform, clone GAD-6 is an antibody against Glutamate Decarboxylase for use in IH & WB.

Key Applications

Immunohistochemistry
Western Blotting

Application Notes

Immunohistochemistry: (<= 1 μg/ml) Optimal working dilutions must be determined by end user.

Immunohystochemical Staining Procedures

The following procedure was developed to localize GAD in rat brain sections of cerebellum. Perform all steps at room temperature unless otherwise indicated. Where normal serum is indicated, use normal serum from the same species as the source of the secondary antibody.This procedure represents suggested guidelines for the use of anti-GAD. Fixation regimen, antibody concentrations, and incubation conditions for a given experimental system should be determined empirically.

1. Perfuse rats with 100 mM phosphate buffer, pH 7.4, containing 1% paraformaldehyde, 0.34% L-lysine, and 0.05% sodium m-periodate (1% PLP).

2. Postfix brains in 1% PLP for 1-2 hours. Longer fixation times may reduce labeling intensity.

3. Transfer brains to 100 mM phosphate buffer containing 30% sucrose, and gently agitate on a shaker platform at +4°C for 48-60 hours.

4. Using a sliding microtome, cut 30 mm sections of frozen cerebellum. As the sections are cut, collect them in a vial of cold 100 mM phosphate buffer.

5. Incubate sections in phosphate-buffered saline (PBS) containing 1.5% normal serum and 0.2% TritonX-100 for 30 minutes.

6. On a shaker platform, incubate sections with anti-GAD (diluted in PBS containing 1.5% normal serum and 0.2% Triton X-100 to a final antibody concentration of 1 mg/ml) for 12-36 hours at +4°C.

7. On a shaker platform, rinse sections eight times, 10-15 minutes per rinse, in PBS.

8. Detect with a standard secondary antibody detection system (Hsu et al., 1981; Falini & Taylor, 1983; Harlow & Lane, 1988; Taylor, 1978).

9. Mount sections, dehydrate, and apply coverslips.

Biological Information
Immunogen	Purified rat brain GAD.
Clone	GAD-6
Concentration	Please refer to the Certificate of Analysis for the lot-specific concentration.
Host	Mouse
Specificity	Recognizes the lower molecular weight isoform of the two GAD isoforms identified in brain (Gottlieb, et al., 1986; Chang & Gottlieb, 1988). This monclonal antibody can be used for immunohistochemical localization in brain or pancreas. Anti-GAD has also been used to label purified GAD on Western blots (Chang & Gottlieb, 1988).
Isotype	IgG2a
Species Reactivity	Human Rat
Antibody Type	Monoclonal Antibody
Entrez Gene Number	NM_000818.1
Entrez Gene Summary	This gene encodes one of several forms of glutamic acid decarboxylase, identified as a major autoantigen in insulin-dependent diabetes. The enzyme encoded is responsible for catalyzing the production of gamma-aminobutyric acid from L-glutamic acid. A pathogenic role for this enzyme has been identified in the human pancreas since it has been identified as an autoantibody and an autoreactive T cell target in insulin-dependent diabetes. This gene may also play a role in the stiff man syndrome.
Gene Symbol	GAD2 MGC161607 GAD65 MGC161605 GAD-65 EC 4.1.1.15
Purification Method	Ammonium sulfate precipitation and DEAE-cellulose chromatography
UniProt Number	Q05329
UniProt Summary	FUNCTION: SwissProt: Q05329 # Catalyzes the production of GABA. COFACTOR: Pyridoxal phosphate. SIZE: 585 amino acids; 65411 Da SUBUNIT: Homodimer (By similarity). SUBCELLULAR LOCATION: Cytoplasm, cytosol. Cytoplasmic vesicle. Cell junction, synapse, presynaptic cell membrane; Lipid-anchor. Golgi apparatus membrane; Peripheral membrane protein; Cytoplasmic side. Note=Associated to cytoplasmic vesicles. In neurons, cytosolic leaflet of Golgi membranes and presynaptic clusters. PTM: Phosphorylated; which does not affect kinetic parameters or subcellular location. & Palmitoylated; which is required for presynaptic clustering. SIMILARITY: SwissProt: Q05329 ## Belongs to the group II decarboxylase family.
Molecular Weight	65 kDa

Biological Information

Immunogen

Purified rat brain GAD.

Clone

GAD-6

Concentration

Please refer to the Certificate of Analysis for the lot-specific concentration.

Host

Mouse

Specificity

Recognizes the lower molecular weight isoform of the two GAD isoforms identified in brain (Gottlieb, et al., 1986; Chang & Gottlieb, 1988). This monclonal antibody can be used for immunohistochemical localization in brain or pancreas. Anti-GAD has also been used to label purified GAD on Western blots (Chang & Gottlieb, 1988).

Isotype

IgG2a

Species Reactivity

Human
Rat

Antibody Type

Monoclonal Antibody

Entrez Gene Number

NM_000818.1

Entrez Gene Summary

This gene encodes one of several forms of glutamic acid decarboxylase, identified as a major autoantigen in insulin-dependent diabetes. The enzyme encoded is responsible for catalyzing the production of gamma-aminobutyric acid from L-glutamic acid. A pathogenic role for this enzyme has been identified in the human pancreas since it has been identified as an autoantibody and an autoreactive T cell target in insulin-dependent diabetes. This gene may also play a role in the stiff man syndrome.

Gene Symbol

GAD2
MGC161607
GAD65
MGC161605
GAD-65
EC 4.1.1.15

Purification Method

Ammonium sulfate precipitation and DEAE-cellulose chromatography

UniProt Number

Q05329

UniProt Summary

FUNCTION: SwissProt: Q05329 # Catalyzes the production of GABA.
COFACTOR: Pyridoxal phosphate.
SIZE: 585 amino acids; 65411 Da
SUBUNIT: Homodimer (By similarity).
SUBCELLULAR LOCATION: Cytoplasm, cytosol. Cytoplasmic vesicle. Cell junction, synapse, presynaptic cell membrane; Lipid-anchor. Golgi apparatus membrane; Peripheral membrane protein; Cytoplasmic side. Note=Associated to cytoplasmic vesicles. In neurons, cytosolic leaflet of Golgi membranes and presynaptic clusters.
PTM: Phosphorylated; which does not affect kinetic parameters or subcellular location. & Palmitoylated; which is required for presynaptic clustering.
SIMILARITY: SwissProt: Q05329 ## Belongs to the group II decarboxylase family.

Molecular Weight

65 kDa

Product Usage Statements
Usage Statement	Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.

Product Usage Statements

Usage Statement

Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.

Storage and Shipping Information
Storage Conditions	Maintain for 1 year at -20°C from date of shipment. Aliquot to avoid repeated freezing and thawing. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.

Storage and Shipping Information

Storage Conditions

Maintain for 1 year at -20°C from date of shipment. Aliquot to avoid repeated freezing and thawing. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.

Packaging Information
Material Size	100 µg

Packaging Information

Material Size

100 µg

实验耗材

分子生物学试剂

蛋白与免疫学

生化试剂

细胞及检测试剂

色谱及层析

仪器设备

实验服务

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Ms X Glutamic Acid Decarboxylase

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