After over 30 years of advances, multiphoton microscopy (MPM) is now instrumental in a wide range of in vivo biological imaging applications. However, it has, until recently, remained not achievable or affordable to meet the unmet need for fast monitoring of biological dynamics and large-scale examination of biological heterogeneity. Only within the past few years have new strategies emerged to empower MPM at a speed that was once inconceivable, notably at kilohertz two-dimensional (2D) frame rate, and 3D rate or beyond. This Review highlights the latest high-speed innovations and discusses the potential of their synergism with other advanced, but less speed-centric MPM toolboxes. Recognizing these prospects and challenges could inspire new approaches for reprioritizing imaging speed in future MPM developments.

经过 30 多年的进步，多光子显微术 (MPM) 现在在广泛的体内生物成像应用中发挥着重要作用。然而，直到最近，它仍然无法实现或负担不起以满足快速监测生物动力学和大规模检查生物异质性的未满足需求。仅在过去几年中，才出现了新的策略，以曾经难以想象的速度增强 MPM，特别是在千赫兹二维 (2D) 帧速率和 3D 速率或更高的速率下。本评论重点介绍了最新的高速创新，并讨论了它们与其他先进但不太以速度为中心的 MPM 工具箱协同作用的潜力。认识到这些前景和挑战可以激发在未来 MPM 发展中重新确定成像速度的新方法。