| Summary: | Understanding the mechanisms and kinetics of membrane damage is of interest to researchers in several overlapping fields of biology. In this study, we describe the development and validation of a simple <inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><msup><mrow></mrow><mn>32</mn></msup></semantics></math></inline-formula>PO<inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><msubsup><mrow></mrow><mn>4</mn><mrow><mn>3</mn><mo>−</mo></mrow></msubsup></semantics></math></inline-formula> release radioassay used to track nanometer-scale damage to the bacterial cell membrane. Nanoscale membrane damage will result in the release of small cytoplasmic molecules, such as amino acids, sugars, and osmolytes. Our radioassay tracks the release of these molecules using the release of cytoplasmic <inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><msup><mrow></mrow><mn>32</mn></msup></semantics></math></inline-formula>PO<inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><msubsup><mrow></mrow><mn>4</mn><mrow><mn>3</mn><mo>−</mo></mrow></msubsup></semantics></math></inline-formula> as a proxy. Our assay can both detect <inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><msup><mrow></mrow><mn>32</mn></msup></semantics></math></inline-formula>PO<inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><msubsup><mrow></mrow><mn>4</mn><mrow><mn>3</mn><mo>−</mo></mrow></msubsup></semantics></math></inline-formula> release and track release kinetics in the order of minutes. We demonstrate the use of our radioassay using <i>A. baumannii</i> treated with colistin and <inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><mrow><mo>Ω</mo><mn>76</mn></mrow></semantics></math></inline-formula>: two agents known to cause membrane damage. Our assay tracks greater membrane damage in <i>A. baumannii</i> treated with both these agents, compared to an untreated control. Our assay fills a niche that is not covered by traditional <inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><msup><mrow></mrow><mn>51</mn></msup></semantics></math></inline-formula>Cr release radioassays and fluorescent staining techniques. Furthermore, our assay can potentially be used to track membrane damage in other membrane systems such as lipid vesicles, animal cells, and organelles.
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