Northern Lights collaboration for better 2-photon probes

描述（由申请人提供）：北极光合作研究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，并具有互补的兴趣和技能，以解决活体大脑活动成像的基本问题。现在大家一致认为，基因编码的荧光生物传感器是唯一适合于提供特异性和多功能性的必要成像大脑中特定细胞的活动。同样清楚的是，要成像厚片细胞或整个大脑，就必须转向双光子（非线性）成像，它能穿透更深入的活体组织，造成更小的损伤。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，人们一直致力于开发更好的荧光蛋白和用于线性成像的生物传感器。不幸的是，由于当前最先进的双光子表征的压倒性复杂性，直接优化双光子非线性特性的探针已经超出了生物传感器工程师的能力范围。这是BRAIN计划的一个关键障碍，因为需要具有高亮度和稳定性的健壮的双光子探针。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器和先进的实验设计开发新工艺和新仪器，这将使参与BRAIN项目的生物传感器工程师能够发展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.