Bioluminescence system assisted by NAD(P)H conversion to increase the sensitivity of quantitative bacterial cell assay

• The conditions of extractants have been studied to ensure sensitivity.
• The bioluminescence of NAD(P)H was used to rapidly enumerate total viable count.
• The content of pyridine nucleotides was determined by bacterial bioluminescence.

The NADH luminescence assay is a rapid, sensitive and easy-to-perform bacterial detection method. However, the detection limit is approximately 107 CFU/mL, which is inadequate for many applications. The purpose of this study is to amplify luminescence assay signals by converting NAD(P)+ to NAD(P)H to provide a more sensitive method for the detection of bacteria. Under optimal conditions, the luminescence intensity correlated well with the bacterial count (R2 = 0.98) and the detection limit was 1.05 × 105 CFU/mL. The sensitivity of this novel bioluminescence enzymatic cycling method is nearly 102 times higher than previous bioluminescence methods. Thus, this improved method can be used to rapidly determine the total viable count with higher sensitivity.Industrial relevanceThis improved method can be used to rapidly determine the total viable count with higher sensitivity in food. This study lays the foundation for the future development of a fast detector using bacterial bioluminescence.