Alanine is a non-essential, non-polar amino acid that has a key metabolic role in the glucose alanine cycle between the liver and tissues. In tissues such as muscle, amino acids are broken down to provide energy; during the process of transamination amino groups are collected in the form of glutamate. Amino groups are then transferred to pyruvate from glutamate by alanine aminotransferase through the glycolysis cycle, which results in the formation of alanine and α-ketoglutarate. Alanine is transported to the liver through the bloodstream. In the liver, alanine aminotransferase performs the reversible reaction consuming alanine and regenerating pyruvate to be used in. Changes in the alanine cycle that increase serum alanine aminotransferase (ALT) levels have been linked to the development of type II diabetes or liver fibrosis in chronic hepatitis B infected patients.

L-Alanine Assay Kit provides a simple, reproducible, and sensitive tool of measuring L-Alanine concentration in plasma, serum, cell lysates, saliva, urine, and other biological liquid samples. In this assay, L-Alanine is decomposed by converter enzyme to generate NADH, which is catalyzed by electron carrier to form a colored product. This product can be read by a microplate reader at 450 nm. Absorbance values are proportional to the L-Alanine levels within the samples.

• Microplate reader capable of measuring absorbance at 450 nm
• Incubator at 37°C
• Precision pipettes to deliver 2 µL to 1 mL volumes
• Tubes to prepare sample dilutions
• 5 mL tubes
• Multi-channel pipettes to deliver 20 µL to 200 µL volumes
• MilliQ water or another type of double distilled water (ddH₂O)

1. Prepare the Reaction Mix, and Control Mix before use.
2. Add 50 µL of the diluted L-Alanine standards or samples to the 96-well microtiter plate.
3. Add 50 µL Reaction Mix to each standard and one half of the paired sample wells, mix thoroughly.
4. Add 50 µL of Control Mix to other half of the paired sample wells. Carefully shake the plate for a few seconds to mix.
5. Incubate the plate for 30 minutes at 37°C with mixing in the dark.
6. Measure the absorbance at 450 nm using a plate reader.