Figure 4

Specificity of the Epic® response. (a) Addition of increasing rHA concentrations to empty wells (green line) or wells containing immobilized bovine fetuin (red line), or asialofetuin (blue line); (b) Addition of rHA to wells containing immobilized human fetuin (purple line), asialofetuin (blue line) or bovine fetuin (red line). Human fetuin and asialofetuin were immobilized at 20 μg/ml; bovine fetuin was immobilized at 150 μg/ml; (c) Addition of native rHA (green line) or rHA that had been heated at 95°C for 5 min (red line) to fetuin immobilized wells; (d) Response of increasing rHA concentrations added to empty wells (green line) or wells containing immobilized bovine fetuin (red line) or asialofetuin (blue line) after a wash and equilibration step; (e) Determination of the linear working rHA concentration range. rHA of B/BR/08 was added to wells containing immobilized fetuin (red line) or asialofetuin (green line) at 5, 10, 40, 80 and 160 μg/ml. In all experiments except those using human fetuin, bovine fetuin and asiaolofetuin were immobilized at 150 μg/ml under optimal conditions. In experiments using human fetuin, immobilization was performed with glycoproteins at 20 μg/ml.