Nori Mouse ALT ELISA Kit Summary

Alternative names for ALT: Alanine transaminase, alanine aminotransferase, ALAT, GPT, GPT1, ALT1, Glutamate pyruvate transaminase 1, Glutamic–alanine transaminase 1, Glutamic–pyruvic transaminase 1

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: 

Alanine transaminase (ALT) is a transaminase enzyme and is also called alanine aminotransferase (ALAT) and was formerly called serum glutamate-pyruvate transaminase (SGPT) or serum glutamic-pyruvic transaminase (SGPT).^[1] ALT is found in plasma and in various body tissues but is most common in the liver. Serum ALT level, serum AST (aspartate transaminase) level, and their ratio (AST/ALT ratio) are commonly measured clinically as biomarkers for liver health. The tests are part of blood panels. ALT catalyzes the transfer of an amino group from L-alanine to α-ketoglutarate, the products of this reversible transamination reaction being pyruvate and L-glutamate. ALT (and all aminotransferases) require the coenzyme pyridoxal phosphate, which is converted into pyridoxamine in the first phase of the reaction, when an amino acid is converted into a keto acid. ALT is commonly measured clinically as part of liver function tests and is a component of the AST/ALT ratio.^[2] Significantly elevated levels of ALT often suggest the existence of other medical problems such as viral hepatitis, diabetes, congestive heart failure, liver damage, bile duct problems, infectious mononucleosis, or myopathy, so ALT is commonly used as a way of screening for liver problems. ^[3] Elevated ALT may also be caused by dietary choline deficiency. However, elevated levels of ALT do not automatically mean that medical problems exist. Fluctuation of ALT levels is normal over the course of the day, and they can also increase in response to strenuous physical exercise. When elevated ALT levels are found in the blood, the possible underlying causes can be further narrowed down by measuring other enzymes. For example, elevated ALT levels due to hepatocyte damage can be distinguished from bile duct problems by measuring alkaline phosphatase. Also, myopathy-related elevations in ALT should be suspected when the aspartate transaminase (AST) is greater than ALT; the possibility of muscle disease causing elevations in liver tests can be further explored by measuring muscle enzymes, including creatine kinase. Many drugs may elevate ALT levels.^[4]

References

1. Karmen A, et al. (1955). The Journal of Clinical Investigation. 34 (1): 126–31.
2. Ghouri N, et al. (2010). Hepatology. 52 (3): 1156–61.
3. Dufour DR, et al. (2000). Clinical Chemistry. 46 (12): 2027–49.
4. Watkins PB, et al. (2006). JAMA. 296 (1): 87–93.