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Analysis Note

ControlRat brain tissue

Application

Detect Glutamate Transporter using this Anti-Glutamate Transporter Antibody, Glial validated for use in IH(P), IF & WB.

Evaluated by Western Blot on mouse brain membrane lysates.

Western Blot: 1:500 dilution of this antibody detected GLT-1 on 10 ug of mouse brain membrane lysates.

Immunohistochemistry:
1:1,000-1:4,000 dilution from a previuos lot used a DAB detection system on adult rat forebrain fixed with 4% paraformaldehyde.

Immunohistochemistry:
1:5,000-10,000 dilution from a previuos lot used a cyanine conjugated secondary antibody.

Enzymatic detection requires substantially higher primary antibody dilutions. Lightly fixed 4% PFA material is recommended.

Notes:

Male Sprague-Dawley rats (b.wt. 100-150g) were anesthetized with sodium pentobarbital and perfused via the ascending aorta with 50 mL of Ca2+-free Tyrode+s solution followed by a formalin-picric acid fixative (4% paraformaldehyde with 0.4% picric acid in 0.16 M phosphate buffer, pH 6.9) for 6 minutes. Tissues were rapidly dissected out, postfixed in the same fixative for 90 minutes and rinsed for at least 24 hours in 0.1 M phosphate buffer (pH 7.4) containing 10% sucrose. Sections were cut (14 um) in a cryostat and incubated at 4°C overnight with AB1783 (1:5,000-1:10,000). After rinsing in PBS sections were incubated for 60 minutes at room temperature with Cy3-conjugated secondary antibodies. After mounting in a mixture of PBS and glycerol (1:3) containing 0.1% p-phenylenediamine, sections were examined with a Nikon Microphot-SA epifluorescence microscope.



Optimal working dilutions must be determined by the end user.

Research Sub CategoryIon Channels & Transporters

Research CategoryNeuroscience

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.

General description

Glutamate transporters (GluT) function to remove L-glutamate (Glu), the primary excitatory neurotransmitter in the mammalian central nervous system (CNS), from the synaptic cleft. By clearing the synapse in this manner Glu can be recycled for later use, the proper diffusion gradient can be maintained and excitotoxicity can be prevented. There have been reports of many different members of a GluT multigene family; GLT1, GLAST, EAAC1 and EAAT2. mRNA has been identified in brain for each of these proteins.

EAAT2 transports L-glutamate and also L- and D-aspartate. It is essential for terminating the postsynaptic action of glutamate by rapidly removing released glutamate from the synaptic cleft. Acts as a symport by cotransporting sodium.

Immunogen

Synthetic peptide from the carboxy-terminus of rat GLT-1.

Epitope: C-terminus

Legal Information

CHEMICON is a registered trademark of Merck KGaA, Darmstadt, Germany

Other Notes

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

Physical form

Guinea pig antiserum containing no preservatives.

Unpurified

Quality

Evaluated by Western Blot on mouse brain membrane lysates.

Specificity

Glial Glutamate Transporter GLT-1 (EAAT2). The antibody has been tested on central nervous system tissue. Preabsorption of the antiserum with the immunogen peptide (Catalog number AG391) completely abolishes the immunostaining.

Storage and Stability

Stable for 1 year at -20°C from date of receipt.

Target description

~62 kDa