Pharmacia phast system

Table 1 indicates the synthesis and characterization of rHDL containing each ketohexose. Discoidal rHDL was prepared by the sodium cholate dialysis method [24 (link)] using initial molar ratios of palmitoyl oleoyl phosphatidylcholine (POPC) : cholesterol : apoA-I : sodium cholate of 95 : 5 : 1 : 150. As the rHDL particles employed in the procedure were pure enough and exhibited extreme uniformity, further processing was deemed unnecessary. PAGGE or native polyacrylamide gradient gel electrophoresis was carried out in order to determine the size of the rHDL particles and to compare them with standard globular proteins (Cat# 17-0445-01 Amersham Pharmacia, Uppsala, Sweden). The Pharmacia Phast system was obtained from GE Healthcare, Uppsala, Sweden. Gel Doc® XR (Bio-Rad, Hercules, CA, USA) complemented with Quantity One software, version 4.5.2 was employed to compare the relative movement of particles, while its content of protein was measured by Lowry method but improvised by as modified Markwell et al. [25 (link)] with standard as BSA.

Isoelectric points for the canola meal crude extracts and purified samples were determined by conducting IEF on the Pharmacia Phast System (GE Healthcare Life Sciences, Uppsala, Sweden). Isoelectric focusing gels termed IEF 3–9 (which are optimized for the pH range of 3–9), were used to resolve proteins based on their isoelectric points. The reference sample used was an Amersham Biosciences broad-range pI calibration kit, and different proteins with well-known isoelectric points ranging from 3 to 10 were used. The protein bands were visualized using the silver staining method as per the manufacturer’s protocol (GE Healthcare Life Sciences, Uppsala, Sweden).