Continuous-flow ATP amplification system for increasing the sensitivity of quantitative bioluminescence assay.
We constructed a novel ATP amplification reactor using a continuous-flow system, and this allowed us to increase the sensitivity of a quantitative bioluminescence assay by controlling the number of ATP amplification cycles. We previously developed a bioluminescence assay coupled with ATP amplification using a batch system. However, it was difficult to control the number of amplification cycles. In this study, ATP amplification was performed using a continuous-flow system, and significant linear correlations between amplified luminescence and initial ATP concentration were observed. When performing four cycles of continuous-flow ATP amplification, the gradient of amplification was 1.87(N). Whereas the lower quantifiable level was 500 pM without amplification, values as low as 50 pM ATP could be measured after amplification. The sensitivity thus increased 10-fold, with further improvements expected with additional amplification cycles. The continuous-flow system thus effectively increased the sensitivity of the quantitative bioluminescence assay.
Satoh
Tetsuya
T
SBI, Interdisciplinary Research on Integration of Semiconductor and Biotechnology, Hiroshima University, Higashi-Hiroshima, Hiroshima 739-8530, Japan.
Shinoda
Yasuharu
Y
Alexandrov
Maxym
M
Kuroda
Akio
A
Murakami
Yuji
Y
D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism*; D000263:Adenylate Kinase / Q000378:metabolism; D004800:Enzymes, Immobilized / Q000737:chemistry / Q000378:metabolism; D008156:Luciferases / Q000737:chemistry / Q000378:metabolism; D008163:Luminescent Measurements / Q000295:instrumentation* / Q000379:methods*; D008954:Models, Biological; D017856:Phosphotransferases (Phosphate Group Acceptor) / Q000378:metabolism; D011770:Pyruvate Kinase / Q000378:metabolism; D015203:Reproducibility of Results
D004800:Enzymes, Immobilized; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D008156:Luciferases; D011770:Pyruvate Kinase; D017856:Phosphotransferases (Phosphate Group Acceptor); C022424:polyphosphate kinase; D000263:Adenylate Kinase