Micro-BIA, a colorimetric microtiter assay of lambda prophage induction

Area of Science:

• Microbiology
• Molecular Biology
• Toxicology

Background:

• Escherichia coli lysogenic for lambda-lacZ fusion phage produce beta-galactosidase upon prophage induction by DNA-damaging agents.
• Assays for detecting DNA-damaging chemicals are crucial for safety and research.

Purpose of the Study:

• To miniaturize a quantitative, colorimetric beta-galactosidase (prophage) induction assay (BIA).
• To develop an efficient screening method for DNA-damaging chemicals.

Main Methods:

• Performed induction assays in microtiter wells using multichannel pipetting devices.
• Utilized screening strain BR513 and an enhanced induction strain BR339.
• Developed a method using frozen, permeabilized bacteria for rapid, consecutive assays.
• Monitored beta-galactosidase levels via colorimetric absorbance or a color chart.
• Generated dose-response curves using various mutagens.

Main Results:

• Demonstrated successful miniaturization and quantification of the beta-galactosidase induction assay.
• Showcased enhanced induction with strain BR339 and a rapid method for its use.
• Validated the assay with a range of chemical mutagens, including polycyclic aromatic hydrocarbons.
• Achieved considerable savings in labor and materials compared to traditional methods.

Conclusions:

• The miniaturized BIA is an effective and economical method for screening DNA-damaging chemicals.
• The assay can be performed with or without automated equipment.
• This assay provides a valuable tool for toxicological screening and research.