Development of a highly sensitive microplate luminometer using ATP bioluminescence

Area of Science:

• Biotechnology
• Analytical Chemistry
• Cell Biology

Background:

• Adenosine Triphosphate (ATP) bioluminescence is crucial for biological assays.
• Existing microplate luminometers have detection limits insufficient for analyzing small cell samples.

Purpose of the Study:

• To develop a highly sensitive, automated microplate luminometer for low ATP concentration analysis.
• To improve detection limits and reduce luminescence crosstalk compared to conventional instruments.

Main Methods:

• Designed an automated system for reagent dispensing and bioluminescence measurement in 96-well microplates.
• Employed a photomultiplier tube (PMT) pressed against the microplate for enhanced light collection and reduced crosstalk.
• Validated performance within 10 minutes for reagent dispensing and measurement.

Main Results:

• Achieved an ATP detection limit of 0.97 amol, an order of magnitude improvement over typical luminometers.
• Reduced luminescence crosstalk to 4.4 × 10^-6, enhancing signal accuracy.
• Demonstrated a wide dynamic range of 10⁶ with consistent linearity across ATP concentrations.

Conclusions:

• The developed automated microplate luminometer offers superior sensitivity and accuracy for ATP detection.
• This novel instrument has broad applicability in biological sciences, particularly for analyzing limited cell samples.
• Enables more precise evaluation of cellular activities and variability at low ATP levels.