Efficient separation and enrichment of pathogenic bacteria from complex matrices are crucial for the detection and downstream biomedical investigations. Herein, we report a floating magnetic membrane comprised of superparamagnetic nanoparticles and cationic polymer chains for rapid capture and enrichment of pathogenic bacteria under continuous flow. Magnetic nanoparticles combined with polymeric chains have shown affordable features to capture, release, and concentrate the pathogens by applying an external magnetic field. We have verified the modulated porous characteristics of the floating magnetic membrane depending on the molecular weight of cationic polymer chains and demonstrated rapid enrichment of pathogenic bacteria from aqueous fluid in the capillary glass tube (> 50-fold). Structural flexibility of the magnetic membrane allows the liquid and smaller species to pass through but efficiently induces binding of the bacteria on the antibody-functionalized magnetic nanoparticles of the floating virtual web. The magnetic membrane enables size-selective filtration and target-specific trapping through ionic exchange and immunomagnetic isolation. This study implies that spatiotemporal application of the magnetic membrane for rapid enrichment of biological targets in a large volume of continuous flow using microfluidic devices and biochips.