Suitable for use in ELISA, Western Blotting, Immuoprecipitation. Other applications not tested.
Recommended Dilution
Western Blot: 1:1000-1:5000 for neat serum and 1-10ug/ml for affinity pure antibody using ECL. This antibody identified a major band at about 52kD in rat liver microsomal membranes (2).
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.
Immuoprecipitation: use at 5-10 ul neat serum per 100ug of rat brain membranes or 1-10ug affinity pure.
Optimal dilutions to be determined by the researcher.
Storage and Stability
May be stored at 4°C for short-term only. For long-term storage and to avoid repeated freezing and thawing, aliquot and add glycerol (40-50%). Freeze at -20°C. Aliquots are stable for at least 12 months at -20°C. 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.
Immunogen
A 12 AA peptide sequence near the C-terminus of rat Glucose Transporter 7 (1). A cysteine has been added at the NH2 terminus for coupling to KLH.
Form
Supplied as a liquid, neat serum, 40% glycerol.
Purity
Serum
Specificity
Rat GT72 sequence is identical in human. Glut-7 and Glut-2 sequences are 68% identical in rats and show similar secondary and tertiary structures. However, Glut-7 contains an extra 6 amino acid (KKMKND) at the C-terminal end. This sequence is very similar to the last 6 amino acids of some of the mircosomal UDP-glucoronosyltransferase isoenzymes (see refs. 1 and other refs. Cited therein). The KK-K-- has been considered motif for the retention of transmembrane proteins (3) and is not found in Gluts 1-5. The crossreactivity of our anti-Glut 7 antibody to other proteins with a endoplasmic reticulum motif and species crossreactivity has not been determined. Antibody crossreactivity in various species is not established. Control peptide, because of its low mol size (<3kD), is not recommended for Western. It should be used in ELISA, Dot Blot and for antibody blocking to confirm specificity of antibodies
References
1. Waddell. I.D., et al (1992) Biochem. J. 286, 173-177 & (1991) Biochem. J. 275, 363-377; 3. Jackson, M.R., et al (1990) EMBO J., 9, 3153-3162