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Filter-based PM_2.5 samples were chemically analyzed to investigate secondary organic aerosol (SOA) formation from isoprene in a rural atmosphere of the southeastern US influenced by both anthropogenic sulfur dioxide (SO₂) and ammonia (NH₃) emissions. Daytime PM_2.5 samples were collected during summer 2010 using conditional sampling approaches based on pre-defined high and low SO₂ or NH₃ thresholds. Known molecular-level tracers for isoprene SOA formation, including 2-methylglyceric acid, 3-methyltetrahydrofuran-3,4-diols, 2-methyltetrols, C₅-alkene triols, dimers, and organosulfate derivatives, were identified and quantified by gas chromatography coupled to electron ionization mass spectrometry (GC/EI-MS) and ultra performance liquid chromatography coupled to electrospray ionization high-resolution quadrupole time-of-flight mass spectrometry (UPLC/ESI-HR-Q-TOFMS). Mass concentrations of six isoprene low-NO_x SOA tracers contributed to 12–19% of total organic matter (OM) in PM_2.5 samples collected during the sampling period, indicating the importance of the hydroxyl radical (OH)-initiated oxidation (so-called photooxidation) of isoprene under low-NO_x conditions that lead to SOA formation through reactive uptake of gaseous isoprene epoxydiols (IEPOX) in this region. The contribution of the IEPOX-derived SOA tracers to total organic matter was enhanced by 1.4% (<i>p</i> = 0.012) under high-SO₂ sampling scenarios, although only weak associations between aerosol acidity and mass of IEPOX SOA tracers were observed. This suggests that IEPOX-derived SOA formation might be modulated by other factors simultaneously, rather than only aerosol acidity. No clear associations between isoprene SOA formation and high or low NH₃ conditional samples were found. Positive correlations between sulfate aerosol loadings and IEPOX-derived SOA tracers for samples collected under all conditions indicates that sulfate aerosol could be a surrogate for surface accommodation in the uptake of IEPOX onto preexisting aerosols.