Northern Lights collaboration for better 2-photon probes

描述(申请人提供)：北极光合作更好的双光子探测器将结合来自蒙大拿州和加拿大的四个非常不同的研究人员和实验室的专业知识，以及互补的兴趣和技能，以解决活着的大脑活动成像的基本问题。现在人们普遍认为，基因编码的荧光生物传感器是唯一适合于提供成像大脑中特定细胞活动所需的特异性和多功能性的生物传感器。同样清楚的是，为了对厚片或整个大脑中的细胞进行成像，有必要转向双光子(非线性)成像，这种成像可以更深入地渗透到活组织中，造成的损害更小。此外，双光子信号提供了单光子信号所缺乏的额外的分子水平信息。多年来，人们一直在开发更好的荧光蛋白质和生物传感器，用于线性成像。不幸的是，由于当前最先进的双光子表征的压倒性的复杂性，直接优化双光子非线性特性的探测器已经超出了生物传感器工程师的能力范围。这是大脑主动性的关键障碍，因为需要具有高亮度和稳定性的强大的双光子探测器。在这里，我们描述了一个为期三年的计划，利用现有的超快激光和先进的实验设计来开发新的工艺和新的仪器，这将使参与大脑项目的生物传感器工程师能够进化出下一代多光子探测器。

DESCRIPTION (provided by applicant): The Northern Lights collaboration for better 2-photon probes will combine the expertise of four very different investigators and laboratories from Montana and Canada with complementary interests and skills to tackle a fundamental problem in imaging of the activity of the living brain. It is now well agreed that genetically encoded, fluorescent biosensors are uniquely suited to provide the specificity and versatility necessary to image the activity of defined cells in the brain. It is also clear that to image cells in thick slies or whole brains it will be necessary to turn to 2- photon (nonlinear) imaging, which penetrates deeper into living tissues and causes less damage. Furthermore, 2-photon signals provide additional molecular-level information that 1- photon signals lack. Many years have gone into developing better fluorescent proteins and biosensors for linear imaging. Unfortunately, directly optimizing probes for the 2-photon nonlinear properties has been beyond the reach of biosensor engineers because of the overwhelming complexity of the current state-of-the-art 2-photon characterization. This is a critical impediment for the BRAIN initiative, because robust 2-photon probes with high brightness and stability are needed. Here we describe a three year program to develop a new process and new instruments using available ultrafast lasers and advanced experimental designs that will empower biosensor engineers involved in the BRAIN project to evolve the next generation of multi photon probes.