Abstract View
Development of a Semi-Automated Instrument to Measure the Cellular Reactive Oxygen Species (Ros) Activity of Ambient Particulate Matter
SUDHEER SALANA, Vishal Verma, University of Illinois Urbana-Champaign
Abstract Number: 73
Working Group: Health-Related Aerosols
Abstract
Several automated instruments exist today to measure the acellular oxidative potential of ambient particulate matter (PM). Moreover, a few online instruments have also been developed for the real time measurements of acellular reactive oxygen species (ROS) activity of the ambient particles. However, no such automated system exists for measuring the cellular ROS activity, which severely limits the comparison between two types of assays. Cellular assays provide a much better means of ROS assessment as they incorporate the biological processes involved in the PM-induced ROS generation. Considering this need, here we aim to develop a semi-automated instrument to conduct the macrophage ROS assay using rat alveolar cells (NR8383), which is a well-established and widely used cell line for the cellular oxidative potential measurements. The instrument uses dichlorofluorescein (DCFH) as a probe to detect the ROS activity of particulate matter. The instrument is capable of analyzing at least two ambient PM samples extracted in water, within a span of four hours, including both negative and positive controls. Preliminary experiments conducted on various standard metal solutions show a very good agreement between manual and automated results for the relative ROS response as compared to the negative control (slope = 0.74 and r2=0.9). The instrument also has a very low limit of detection (<2.6 µg equivalents of Zymosan). The analysis of the time dependent ROS response, and measurement of the ROS activity of the PM-laden filter samples collected during field trials is currently underway, which will help to further evaluate the precision and accuracy of the system. Coupled with other acellular assays, this instrument would provide a comprehensive analysis of the cellular oxidative potential of particular matter samples.