Anti horse spleen ferritin antibody

Proteins were dialyzed overnight in Buffer A: 100mM Tris HCl, pH 8.0 at 37°C, followed by centrifugation for 30 min at 8,000x g at 4°C to remove protein aggregates. Protein concentration was determined using A260/280 or the Bradford method.^{15 (link)} Kinetics of protein aggregation were determined by measuring of apparent absorption due to light scattering at 360nm (A360) in a DU-650 spectrophotometer (Beckman, USA) equipped with a Peltier temperature controller. Substrates and chaperone-like proteins were mixed in 400μL black walled cuvettes at molar ratios as described in the text. The scattering in each cell was recorded automatically each 1 min over a 1 hour interval at fixed temperature (37°C and 43°C). For analysis of heat treated precipitates, samples were incubated at 43°C for 15 min, centrifuged at 10Kg × 30 min, the precipitates were dissolved in 4% SDS and subjected to SDS-PAGE and Western blot analysis using a polyclonal rabbit antiserum to CS purchased from Nordic Immunological Laboratories (Tiburg, Netherlands) and anti-horse spleen ferritin antibody developed in rabbit (Sigma-Aldrich, Saint Louis, USA).