Antiviral activities of SAP and PTX3 against IAV and IAV-infected cells.

<p>(<b>A</b>) Inhibition of NA activity by SAP and PTX3. A dilution of HKx31 was mixed with a final concentration of 10 or 1 µg/ml of SAP (white bars) or PTX3 (black bars) in BSA₅-TBS-Ca²⁺ and assayed for NA activity as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0059623#s2" target="_blank">Materials and Methods</a>. Results are expressed as the percent of NA activity compared to a virus only control. Data shown are the mean (±1 SD) from three independent experiments. *p<0.05, Students <i>t</i>-test, significant difference in levels of NA activity in the presence of SAP compared to an equivalent concentration of PTX3. (<b>B</b>) Neutralization of IAV by SAP and PTX3. Pentraxins were treated with sialidase from <i>Vibrio cholerae</i> (50 mU) in TBS containing 10 mM Ca²⁺ or mock-treated (M) in buffer alone for 30 min at 37°C, and then heated to 56°C for 30 min to inactivate sialidase. Sialidase- (S) or mock- (M) treated pentraxins were mixed with a dilution of HKx31 to give a final concentration of 10 or 1 µg/ml of PTX3 or SAP, and the amount of infectious virus remaining determined by fluorescent-focus reduction assay. Results are expressed as a percent of the number of fluorescent foci in the virus only control. Data show the mean (±1 SD) from three independent experiments. *p<0.05, ***p<0.001, Student’s <i>t</i>-test, significant difference between sialidase-treated sample compared to appropriate mock-treated control. (<b>C</b>) Binding of SAP and PTX3 to IAV-infected MDCK cells. Binding of SAP and PTX3 to uninfected (i, iii) or HKx31-infected MDCK cells (ii, iv) was determined at 6 hrs post-infection. Aliquots of 10⁶ cells were incubated with biotin-labelled SAP (10 µg/ml) or PTX3 (2 µg/ml) in BSA₅-TBS-Ca²⁺ (white histograms) or BSA₅-TBS-EDTA (grey histograms). Binding of biotin-labelled pentraxins was determined by flow-cytometry. Histograms are representative of two independent experiments.