Using horseradish peroxidase (HRP)-HDL3, instead of the classical 125 I-HDL3, an enzyme linked assay for measuring HDL recptor on cultured human arterial smooth muscle cells (SMC )was developed. Horseradish peroxidase...Using horseradish peroxidase (HRP)-HDL3, instead of the classical 125 I-HDL3, an enzyme linked assay for measuring HDL recptor on cultured human arterial smooth muscle cells (SMC )was developed. Horseradish peroxidase labeled HDI3, was prepared by periodate method.Human arterial SMC were fixed on 96-wells plastic plates by glutaraldehyde mehtod. The results showed that Scatchard plot of high affinity binding of high affinity binding of HRP-HDL3 to SMC was significantly linear,r=—0.987. The Kd and Bmax were 13.12±0.8μg/ml and 106±9.3 ng/mg cell protein, respectively. HDL3 competed effectively for binding of HRP-HDL3 to SMC. On the contrary,LDL and albumin did not compete with HRP-HDL binding. The binding was Ca2+ independent and not sensitive to trypsin. These results indieated that human arterial smooth muscle cells possess specific binding sites for HDL with high affinity.展开更多
文摘Using horseradish peroxidase (HRP)-HDL3, instead of the classical 125 I-HDL3, an enzyme linked assay for measuring HDL recptor on cultured human arterial smooth muscle cells (SMC )was developed. Horseradish peroxidase labeled HDI3, was prepared by periodate method.Human arterial SMC were fixed on 96-wells plastic plates by glutaraldehyde mehtod. The results showed that Scatchard plot of high affinity binding of high affinity binding of HRP-HDL3 to SMC was significantly linear,r=—0.987. The Kd and Bmax were 13.12±0.8μg/ml and 106±9.3 ng/mg cell protein, respectively. HDL3 competed effectively for binding of HRP-HDL3 to SMC. On the contrary,LDL and albumin did not compete with HRP-HDL binding. The binding was Ca2+ independent and not sensitive to trypsin. These results indieated that human arterial smooth muscle cells possess specific binding sites for HDL with high affinity.
