This study presents a method for rapid, three-dimensional (3D) isotropic imaging of living cells using Bessel beam plane illumination. The technique uses scanned Bessel beams in conjunction with structured illumination and/or two-photon excitation to generate thin light sheets (<0.5 μm) suitable for 3D subcellular imaging. The microscope achieves 3D isotropic resolution down to ~0.3 μm, speeds up to nearly 200 image planes per second, and allows noninvasive acquisition of hundreds of 3D data volumes from single living cells, encompassing tens of thousands of image frames. The method improves upon traditional plane-illumination microscopy by reducing out-of-focus excitation and enhancing axial resolution. It uses a Bessel beam to create a thin light sheet, which is scanned across the focal plane of the detection objective. This allows for high-speed volumetric imaging without moving the specimen. The technique also incorporates structured illumination (SI) to further improve axial resolution and reduce background noise. The study compares the performance of the Bessel beam plane illumination microscope with confocal and digital scanned laser light sheet (DSLM) microscopes. The Bessel beam method achieves superior axial resolution and reduced photobleaching and phototoxicity, enabling extended observations of living cells with isotropic resolution at high volumetric frame rates. It is particularly effective for imaging dynamic cellular processes such as mitochondrial dynamics, filopodia, membrane ruffles, and mitotic chromosomes. The method also supports two-photon excitation, which suppresses Bessel side lobes and allows for a thinner light sheet. This results in high axial resolution and minimal out-of-focus excitation. The Bessel beam method is well-suited for live-cell imaging due to its reduced phototoxicity and ability to capture long-term dynamic processes. The study demonstrates the effectiveness of the Bessel beam plane illumination microscope in achieving high-resolution, high-speed 3D imaging of living cells. It offers a powerful tool for studying cellular dynamics and subcellular structures in living systems. The method is particularly useful for imaging thick, densely fluorescent specimens and has potential applications in live-cell imaging, 3D anatomical mapping, and in vivo imaging of neuronal activity.This study presents a method for rapid, three-dimensional (3D) isotropic imaging of living cells using Bessel beam plane illumination. The technique uses scanned Bessel beams in conjunction with structured illumination and/or two-photon excitation to generate thin light sheets (<0.5 μm) suitable for 3D subcellular imaging. The microscope achieves 3D isotropic resolution down to ~0.3 μm, speeds up to nearly 200 image planes per second, and allows noninvasive acquisition of hundreds of 3D data volumes from single living cells, encompassing tens of thousands of image frames. The method improves upon traditional plane-illumination microscopy by reducing out-of-focus excitation and enhancing axial resolution. It uses a Bessel beam to create a thin light sheet, which is scanned across the focal plane of the detection objective. This allows for high-speed volumetric imaging without moving the specimen. The technique also incorporates structured illumination (SI) to further improve axial resolution and reduce background noise. The study compares the performance of the Bessel beam plane illumination microscope with confocal and digital scanned laser light sheet (DSLM) microscopes. The Bessel beam method achieves superior axial resolution and reduced photobleaching and phototoxicity, enabling extended observations of living cells with isotropic resolution at high volumetric frame rates. It is particularly effective for imaging dynamic cellular processes such as mitochondrial dynamics, filopodia, membrane ruffles, and mitotic chromosomes. The method also supports two-photon excitation, which suppresses Bessel side lobes and allows for a thinner light sheet. This results in high axial resolution and minimal out-of-focus excitation. The Bessel beam method is well-suited for live-cell imaging due to its reduced phototoxicity and ability to capture long-term dynamic processes. The study demonstrates the effectiveness of the Bessel beam plane illumination microscope in achieving high-resolution, high-speed 3D imaging of living cells. It offers a powerful tool for studying cellular dynamics and subcellular structures in living systems. The method is particularly useful for imaging thick, densely fluorescent specimens and has potential applications in live-cell imaging, 3D anatomical mapping, and in vivo imaging of neuronal activity.