The biochemical performance of D-luciferin is assayed at 25°C by an automatic procedure using a Berthold Luminometer. The assay is based on standardized preparations of all components (luciferase, ATP, buffer etc.) required for the measurement. The light emission values obtained with 1.000 and 0.250 mg/mL of D-Luciferin and compared with a reference lot of D-Luciferin.

Actual results for 5 samples from a typical lot, in the absence of substrate inhibition and non-competitive inhibitors, will approach or exceed 100%. With test and reference lots of similar qualities a value around 100% is expected.

Table 1: Biochemical performance of D-Luciferin

Reference is used in concentration of 0.200 mg/mL	Test 0.200 mg/mL
Lot No.1021	101.0 ± 1.9%
Specification	> 97.0%

DISSOLVING D-Luciferin

Warm Luciferin substrate reagent to room temperature before starting.

D-luciferin (acid form) can be dissolved at a concentration of 10 mg/mL (35.7 mmol/L) in an equimolar concentration of base (pH 7.6 to 7.8), e. g. Tris -Tricine and Tris EDTA and HEPES buffers.

A solution is readily obtained using stoichiometric amounts of diluted NaHCO₃ solution. Add 100 mg Luciferin into 3 mL of 1% NaHCO₃ in a buffer. Vortex to obtain immediately a clear yellow solution which should be protected from direct light exposure.