Fig 1. Inactivation of MN inhibitory activity with a heat treatment step.

Human serum and plasma that was spiked with calcium chloride and the indicated amounts of MN was either heated to 90°C or incubated at 4°C, for 20 minutes (A). Unheated samples and supernatants of heated samples were then incubated with the 11mer PolyT FAM probe and fluorescence was measured. Note the clear differences between the heated and unheated samples with MN concentrations below 10⁻² units/μl (panel in A). Error bars indicate standard deviations of triplicate measurements. The incubation time needed to inactivate inhibitory antibodies in plasma at 90 and 100°C was determined as follows. Human plasma, spiked with 10⁻⁵ units/μl MN and 10 mM calcium chloride, was heated to 90°C (upper panel in B) or 100°C (lower panel in B) for indicated amounts of time, centrifuged, and supernatants were incubated with the 11mer polyT FAM probe. Probe incubated with buffer only was included as control. Error bars indicate standard deviations of triplicate measurements.