Technical Data
E0255-05B
EAT-4 (Vesicular Type Transporters for Glutamate, VGLUT)
Description:
Glutamate is the main excitatory neurotransmitter in the brain. To date five glutamate Transporters have been cloned: GLAST (EAAT1), GLT1 (EAAT2), EAAC1 (EAAT3), EAAT4, and EAAT5. These transporters are believed to be critical in reducing potentially toxic extracellular concentration of glutamate by rapid uptake into nerve terminals and glial cells. Most recently, vesicular type transporters for glutamate, termed VGLUTs (VGLUT1/BNPI, VGLUT2/DNPI, and EAT-4), have been cloned and characterized that are related to phosphate transporters. Although neurons exhibit Na-dependent Pi transport, the biological role of Pi uptake is not clear. Proximal tubules in the kidney reabsorb Pi in the glomeruli by the action of a group of phosphate transporters (Type1-NaPi related, type2-NaPi-2 related, and type 3-viral receptor-related. These receptors show weak (~20% identity) between various subtypes. A distinct type of brain specific Na+-dependent phosphate (Pi) transporter (BNPI), originally characterized as a plasma membrane transporter has been localized in a subset of glutamatergic neurons (amygdla,cereberal cortex, and hippocmaplus) and identified as VGLUT1 (rat and human 560 aa; ~60kD, ~30% homology with type-1 Pi-transporters). VGLUT1 shows strong sequence homology to EAT-4, a C. elegans protein that appears to have specific presynaptioc role in glutamtergic transmission. Loss of function mutations in EAT-4 affects multiple glutamregic neurotransmission pathways. EAT4 encodes a protein of 563 aa (~48% identity with BNPI and ~30% identity with known NaPi-transporters found in mammalian kidney). Like BNPI, EAT-4 is also expressed in glutamatergic neurons.

Applications:
Suitable for use in ELISA, Western Blotting. Other applications not tested.

Recommended Dilution:
Western Blot: 1:1000-1:5000 for antiserum and 1-10ug/ml for affinity pure antibody using Chemiluminescence technique. EAT-4 is ~65kD protein.
ELISA: 1:10,000-50,000 for neat serum and 0.5-1ug/ml for affinity pure. Control peptide can be used to coat ELISA plates at 1ug/ml.

Optimal dilutions to be determined by the researcher.

Storage and Stability:
May be stored at 4C for short-term only. For long-term storage and to avoid repeated freezing and thawing, aliquot and add glycerol (40-50%). Freeze at -20C. Aliquots are stable for at least 12 months at -20C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.
TypeIsotypeCloneGrade
PabIgGSerum
SizeStorageShippingSourceHost
100ul-20CBlue IceRatRabbit
Concentration:
As reported
Immunogen:
An 18 AA synthetic peptide witin the N-terminus of C. elegans EAT-4 was synthesized, conjugated to KLH
Purity:
Serum
Form
Supplied as a liquid, neat serum, 40% glycerol.
Specificity:
The peptide has no significant sequence with mammalian VGLUT1/2 or other NaPi-related (Type-1 to Type-III) transporters. Antibody crossreactivity in various species is not established. Control peptide, because of its low MW (<3kD), is not suitable for Western. It should be used for ELISA or antibody blocking (use 5-10 ug per 1 ul of antiserum or 1 ug of aff pure IgG) to confirm antibody specificity.
Intended for research use only. Not for use in human, therapeutic, or diagnostic applications.
1. Lee RYN et al (1999) J. Neurosci. 19:159-167