Comparison of Picogreen and Sytox Orange Stains in Quantitative Analysis of Extracellular Trap Formed against Toxoplasma gondii in Polymorphonuclear Leukocytes from Sheep.

Abstract

OBJECTIVE: The present study aimed to compare the effectiveness of two extracellular DNA dyes (Sytox Orange and PicoGreen) in quantitative measurement of the DNA that forms the backbone of the extracellular trap structures in vitro. Toward this aim, the co-culture of polymorphonuclear leucocytes (PMNs) and Toxoplasma gondii tachyzoites isolated from sheep was selected as model. METHODS: T. gondii tachyzoites and PMNs isolated from the sheep were incubated for varied durations (30, 60, 90 and 120 min); the extracellular DNA released following this incubation was stained using two different dyes. RESULTS: In the present study, no statistical significant difference was observed between the two extracellular DNA dyes with regard to the measurement of extracellular DNA released for all the incubation durations (p>0.05), except for the 30-min incubation (p=0.014). CONCLUSION: There was a statistically significant difference at 30 min incubation time. This difference may be attributed to the staining properties of the dye. Researchers studying in vitro netosis are recommended to use two different extracellular DNA dyes for the quantitative analysis of extracellular DNA formed during short-term incubation.