Nori Human ACC1 ELISA Kit Summary

Alternative names for ACC1: Acetyl-CoA carboxylase 1, Acetyl-CoA carboxylase alpha, ACACA, ACAC, ACCA

Standard Curve

Kit Components
1. Pre-coated 96-well Microplate
2. Biotinylated Detection Antibody
3. Streptavidin-HRP Conjugate
4. Lyophilized Standards
5. TMB One-Step Substrate
6. Stop Solution
7. 20 x PBS
8. Assay Buffer

Other Materials Required but not Provided:
1. Microplate Reader capable of measuring absorption at 450 nm
2. Log-log graph paper or computer and software for ELISA data analysis
3. Precision pipettes (1-1000 µl)
4. Multi-channel pipettes (300 µl)
5. Distilled or deionized water

Protocol Outline
1. Prepare all reagents, samples and standards as instructed in the datasheet.
2. Add 100 µl of Standard or samples to each well and incubate 1 h at RT.
3. Add 100 µl of Working Detection Antibody to each well and incubate 1 h at RT.
4. Add 100 µl of Working Streptavidin-HRP to each well and incubate 20 min at RT.
5. Add 100 µl of Substrate to each well and incubate 5-30 min at RT.
6. Add 50 µl of Stop Solution to each well and read at 450 nm immediately.

Background: 

Acetyl-CoA carboxylase (ACC) is a biotin-dependent enzyme that catalyzes the irreversible carboxylation of acetyl-CoA to produce malonyl-CoA through its two catalytic activities, biotin carboxylase (BC) and carboxyltransferase (CT). The most important function of ACC is to provide the malonyl-CoA substrate for the biosynthesis of fatty acids.^[1] When the enzyme is active, the product, malonyl-CoA, is produced which is a building block for new fatty acids and can inhibit the transfer of the fatty acyl group from acyl CoA to carnitine with carnitine acyltransferase, which inhibits the beta-oxidation of fatty acids in the mitochondria. The activity of ACC can be controlled at the transcriptional level as well as by small molecule modulators and covalent modification. The human genome contains the genes for two different ACCs^[2]—ACACA^[3] and ACACB.^[4] In mammals where two isoforms of ACC are expressed, the main structural difference between these isoforms is the extended ACC2 N-terminus containing a mitochondria targeting sequence.^[1] ACC1 is found in the cytoplasm of all cells but is enriched in lipogenic tissue, such as adipose tissue and lactating mammary glands, where fatty acid synthesis is important.^[5] In oxidative tissues, such as the skeletal muscle and the heart, the ratio of ACC2 expressed is higher. ACC1 and ACC2 are both highly expressed in the liver where both fatty acid oxidation and synthesis are important.^[6] The differences in tissue distribution indicate that ACC1 maintains regulation of fatty acid synthesis whereas ACC2 mainly regulates fatty acid oxidation. Mammalian ACC1 and ACC2 are regulated transcriptionally by multiple promoters which mediate ACC abundance in response to the cells nutritional status. A lack of ACC1 in mutant mice is lethal already at the embryonic stage. ACC2 -/- mice were protected from diabetes^[7] and have continuous fatty acid oxidation, reduced body fat mass, and reduced body weight despite an increase in food consumption.

References

1. Tong L (2005). Cell. Mol. Life Sci. 62 (16): 1784–803.
2. Brownsey RW, et al. (1997). Biochem. Soc. Trans. 25 (4): 1232–8.
3. Abu-Elheiga L, et al. (1995). Proc. Natl. Acad. Sci. U.S.A. 92 (9): 4011–5.
4. Widmer, J; et al. (1996). J. 316 (3): 915–22. doi:10.1042/bj3160915.
5. Kim TS, et al. (1996). Biochem. Biophys. Res. Commun. 225 (2): 647–53.
6. Barber MC, et al. (2005). Biochim. Biophys. Acta. 1733 (1): 1–28.
7. L Abu-Elheiga; et al. (2001). Science. 291 (5513): 2613–6.