The branched-chain amino acids or BCAAs, refer to the amino acids with non-linear aliphatic side-chains, namely leucine, isoleucine and valine. The BCAA may function in both energy production and as nutrient signals. Dietary supplementation with BCAA may aid in the treatment of various disorders and to prevent muscle atrophy in cancer patients. BCAA may increase the rate of protein synthesis and sensitize cells to insulin and insulin-like growth factor. BCAAs are currently used clinically to aid in the recovery of burn victims, as well as for strength supplementation for athletes. BCAAs, primarily Leu, can stimulate insulin secretion. The BCAAs have also been implicated in a wide range of other physiological effects.

Branched Chain Amino Acid Assay Kit provides a simple, reproducible, and sensitive tool of measuring BCAA concentration in plasma, serum, cell lysates, saliva, urine, and other biological liquid samples. This assay employs a coupled enzymatic reaction system that combines multiple steps. Initially, a BCAA molecule is converted to its ketone form, which results in the conversion of NAD+ to NADH. NADH is then detected by a colorimetric indicator, with an absorption maximum of 450 nm. The optical density measured at 450 nm is directly proportional to the concentration of BCAA present in the sample.

• Microplate reader capable of measuring absorbance at 450 nm
• 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-Leucine 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 room temperature with mixing in the dark.
6. Measure the absorbance at 450 nm using a plate reader.