A membrane immuno-chromatographic system that selectively separates plasma lipoproteins and generates a signal in proportion to the concentration of cholesterol (HDL-C) within high-density lipoprotein (HDL) was investigated as a point-of-care device for the prognosis of coronary heart disease. The system consists of three functional membrane strip pads connected in a sequence for: (from the bottom) immuno-separation based on biotinstreptavidin reaction, catalytic conversion of cholesterol to hydrogen peroxide, and production of a signal. For immuno-chromatography, a monoclonal antibody, specific to apolipoprotein B100 that is present on the surfaces of low-density lipoproteins (LDL) and very low-density lipoproteins (VLDL), with a high binding constant (5×10(10)L/mol) was raised and chemically conjugated to streptavidin. The conjugate was first reacted with lipoprotein particles, and this mixture was absorbed by the capillary action into the biotin pad of the system. After being transferred by medium, immuno-capture of LDL and VLDL particles onto the biotin pad took place, and in situ generation of a signal in proportion to HDL-C consecutively occurred. The capture was selective as well as effective (minimum 90% of LDL and VLDL in clinical concentration ranges), and the detection limit of HDL-C was far lower than 20 mg/dL. To construct a user-friendly device, we are currently investigating the automation of such processes of reactions and separation by adapting a liquid flow-controlling technology that programs the times for the immune reaction and separation. My group further pursues an interdisciplinary study to develop a micro system employing semiconductor-based technologies that will eventually enable the handling of sub-micro liter volume of body fluid as a specimen.

Point-of-Care-Testing Membrane Strip Immuno-Chromatography Plasma Lipoprotein Cholesterol Non-Invasive Diagnosis

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