Fluorescence analysis of serum lipoproteins using a histidine-imidazole polyacrylamide slab gel electrophoresis method
Yasuhiro Takenaka, Ikuo Inoue, Masaaki Ikeda, Yoshihiko Kakinuma · bioRxiv (Cold Spring Harbor Laboratory) · 2025
Abstract Polyacrylamide gel electrophoresis (PAGE) has traditionally been used to analyze lipoproteins in human and animal sera. Most conventional PAGE systems for lipoprotein analysis use disc electrophoresis, which limits the number of samples simultaneously analyzed and does not allow the comparison of electrophoretic profiles of multiple samples under the same conditions. Other methods, such as high performance liquid chromatography and agarose gel electrophoresis, are also used for lipoprotein analysis; however, they require specialized equipment and expertise. Here, we report an improved slab PAGE method with high-throughput, cost-effective, simple, rapid, and reproducible analysis of lipoproteins in the human serum. This system uses Tris-histidine as the running buffer and imidazole buffer for the polyacrylamide gel; therefore we named it the histidine-imidazole polyacrylamide gel electrophoresis (HI-PAGE) system. HI-PAGE effectively prevented band distortions of lipoproteins within an hour of electrophoresis and guaranteed precise quantitative analyses of multiple samples containing lipid-protein complexes. Furthermore, we pre-stained the lipoproteins with a fluorescent dye, Nile Red, and applied it to HI-PAGE electrophoresis. Fluorescent HI-PAGE (fHI-PAGE) was applied to the clinical samples and revealed that the method is highly sensitive and allows the quantitative detection of lipoproteins in human serum.