MAB351
Anti-Glutamate Decarboxylase Antibody, 65 kDa isoform, clone GAD-6
clone GAD-6, Chemicon®, from mouse
Synonym(s):
GAD65
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About This Item
UNSPSC Code:
12352203
eCl@ss:
32160702
NACRES:
NA.41
biological source
mouse
Quality Level
antibody form
purified immunoglobulin
antibody product type
primary antibodies
clone
GAD-6, monoclonal
species reactivity
rat, human
manufacturer/tradename
Chemicon®
technique(s)
immunohistochemistry: suitable
western blot: suitable
isotype
IgG2a
NCBI accession no.
UniProt accession no.
shipped in
dry ice
target post-translational modification
unmodified
Gene Information
human ... GAD2(2572)
General description
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 (65 kDa and 67 kDa, 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 (585 a.a.), encoded on human chromosome 10, and is responsible for vesicular GABA production. GAD67 is cytoplasmic (594 a.a.), 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.
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).
Immunogen
Purified rat brain glutamic acid decarboxylase.
Application
Anti-GAD Antibody. 65 kDa isoform, clone GAD-6 detects level of Glutamate Decarboxylase & has been published & validated for use in IH & WB.
Immunohistochemistry: ( 1 μg/mL * See protocol)
Western blot
Optimal working dilutions must be determined by end user.
APPLICATION NOTES FOR MAB351
IMMUNOHISTOCHEMISTRY
1) Perfuse rats with 100 mM phosphate buffer, pH 7.4 containing 1% paraformaldehyde, 0.34% L-lysine and 0.05% m-periodate (1% PLP).
2) Postfix brains in 1% PLP for 1-2 hours.
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 PBS containing 1.5% normal serum and 0.2% Triton X-100 for 30 minutes.
6) On a shaker platform, incubate sections with MAB351 (diluted 1 μg/mL in PBS containing 1.5% normal serum and 0.2% Triton X-100) 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 standard secondary antibody detection system (PAP, ABC, etc.).
9) Mount sections, dehydrate, and apply coverslips.
Western blot
Optimal working dilutions must be determined by end user.
APPLICATION NOTES FOR MAB351
IMMUNOHISTOCHEMISTRY
1) Perfuse rats with 100 mM phosphate buffer, pH 7.4 containing 1% paraformaldehyde, 0.34% L-lysine and 0.05% m-periodate (1% PLP).
2) Postfix brains in 1% PLP for 1-2 hours.
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 PBS containing 1.5% normal serum and 0.2% Triton X-100 for 30 minutes.
6) On a shaker platform, incubate sections with MAB351 (diluted 1 μg/mL in PBS containing 1.5% normal serum and 0.2% Triton X-100) 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 standard secondary antibody detection system (PAP, ABC, etc.).
9) Mount sections, dehydrate, and apply coverslips.
Research Category
Neuroscience
Neuroscience
Research Sub Category
Neurotransmitters & Receptors
Neurotransmitters & Receptors
Target description
65 kDa
Physical form
Format: Purified
Protein A purified
Purified immunoglobulin. Lyophilized from 10 mM potassium phosphate, 70 mM sodium chloride, pH 7.4, 0.02% sodium azide. Reconstitute with 100 μL of sterile water (1 mg/mL).
Storage and Stability
Maintain lyophilized material at -20°C for up to 12 months. After reconstitution maintain frozen at -20°C in undiluted aliquots for up to 6 months. Avoid repeated freeze/thaw cycles.
Analysis Note
Control
Rat brain tissue
human brain lysate
Rat brain tissue
human brain lysate
Other Notes
Concentration: Please refer to the Certificate of Analysis for the lot-specific concentration.
Legal Information
CHEMICON is a registered trademark of Merck KGaA, Darmstadt, Germany
Disclaimer
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.
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Signal Word
Warning
Hazard Statements
Precautionary Statements
Hazard Classifications
Acute Tox. 4 Dermal - Acute Tox. 4 Inhalation - Acute Tox. 4 Oral - Aquatic Chronic 3
Storage Class Code
13 - Non Combustible Solids
WGK
WGK 3
Flash Point(F)
Not applicable
Flash Point(C)
Not applicable
Certificates of Analysis (COA)
Search for Certificates of Analysis (COA) by entering the products Lot/Batch Number. Lot and Batch Numbers can be found on a product’s label following the words ‘Lot’ or ‘Batch’.
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