BioSMART: BIOreactor Spatial Mapping and Actuation in Real Time

我们今天拥有的许多药物和化学品不是在化工厂生产的，而是在生物反应器中生产的：含有微生物或其他细胞的容器可以产生我们想要的化学物质。我们这样做是因为使用活细胞来制造复杂的化学品在能源，原材料和制造工厂本身的成本方面可以便宜得多（不需要像一些化工厂那样在高温高压下运行）。然而，生物反应器比它们的化工厂表亲更难使用，因为活细胞对它们的环境非常敏感，而且有一系列复杂且难以建模的方式。确保生物反应器以最大能力工作的唯一明智的方法是观察反应过程中发生了什么。在这个项目中，我们希望第一次通过使用细胞本身来监测整个生物反应器中细胞的状况，以告诉我们发生了什么。我们可以通过遗传修饰细胞来根据其局部环境改变其物理特性。我们称这些转基因细胞为生物传感器，在我们的研究中，它们通过荧光报告自己的状态：制造一种蛋白质，当你照射它时，它会发光。虽然研究已经证明了生物传感器的发展及其在生物加工中的好处，但到目前为止，生物传感器在工业过程中的应用一直受到缺乏基础设施的阻碍。这是因为迄今为止大多数分析技术都不是活的，而是基于信号的化学或物理发展。为了真正利用生物传感器增强的灵敏度和特异性，需要开发硬件和数据分析工具，将其集成到工业生物反应器中。这项提议试图填补这一空白。监测荧光是一个挑战;生物反应器本身并不漂亮和透明，而是阴暗和浑浊。我们不能只是透过它看，因为来自外部的光在再次射出之前会散射（或反弹）多次。幸运的是，有一种称为荧光漫射光学断层扫描（fDOT）的技术可以解释散射光。它不能像显微镜一样分辨小的特征，但在生物反应器中这并不重要，因为厘米级的分辨率就足够了。我们将建立一个可以使用fDOT监控整个生物反应器的系统，通过在反应器表面的不同点照射激光并观察从各个侧面的细胞产生的辉光;通过从许多不同的位置进行测量并使用合适的计算机算法，我们可以得到发光细胞如何分布的3D模型。有了这些信息，我们就可以使用建模来预测细胞行为，并自动控制生物反应器的条件，以提高产量。作为演示，我们将重点监测乳酸的积累，乳酸是厌氧发酵的副产品。（贫氧）反应条件;过量的乳酸是有毒的，并且可以限制（甚至杀死）产生乳酸的细胞。通过设计细胞根据附近乳酸的多少发光，我们可以监控反应，或者增加加入到反应中的氧气量，搅拌罐，或者甚至重新设计反应器本身以避免反应条件的局部差异。乳酸的情况只是一个例子;未来的细胞可能会报告温度、剪切力、氧合或任何其他参数的变化，每个参数都有不同颜色的发光。总的来说，这个项目代表了迈向新前沿的第一步，在生物反应器中的细胞不仅能产生我们想要的化学物质，但是告诉我们反应中出了什么问题以及如何修复它。结果是一个反应堆可以更便宜地制造复杂的化学品，考虑到我们现代世界对这些化学品的依赖程度，这对全球经济有着微妙但明显的好处。

Many drugs and chemicals we have today are made not in chemical plants, but in bioreactors: vessels containing microbes or other cells which can create the chemical we want. We do this because using living cells to create complex chemicals can be much cheaper in terms of energy, raw materials and the cost of making the plant itself (which does not need to operate at high temperature and pressure as some chemical plants do). Nevertheless, bioreactors are harder to use than their chemical plant cousins, because living cells are sensitive to their environment in a range of complex and difficult-to-model ways. The only sensible way to make sure that the bioreactor is working at maximum capacity is to watch what is going on during the reaction.In this project we want to, for the first time, monitor the conditions of cells throughout the bioreactor by using the cells themselves to tell us what is what is going on. We can do this by genetically modifying the cells to change their physical properties based on their local environment. We call these genetically-modified cells biosensors, and in our case they report their condition by fluorescence: making a protein which glows when you shine light on it. While studies have demonstrated the development of biosensors and their benefits in bioprocessing, so far the implementation of biosensors in industrial processes has been hampered by a lack of infrastructure for their use. This is because most analytical techniques to date have not been living, but rather based on chemical or physical development of signals. In order to really capitalise on the enhanced sensitivity and specificity of biosensors, development of hardware and data analysis tools for integrating them into industrial bioreactors is needed. This proposal seeks to fill this gap.Monitoring the fluorescent glow is a challenge; the bioreactor itself isn't nice and transparent, but murky and turbid. We can't just look through it, because light from the outside will scatter (or bounce) multiple times before it gets out again. Fortunately, there is a technique called Fluorescence Diffuse Optical Tomography (fDOT) which can account for scattered light. It cannot resolve as small features as a microscope, but in a bioreactor this isn't important, as centimeter-scale resolution is enough. We will build a system that can monitor the whole bioreactor using fDOT, by shining a laser at different points on the reactor surface and watching the resulting glow from the cells on all sides; by taking measurements from lots of different locations and using a suitable computer algorithm, we can get a 3D model of how the glowing cells are distributed. With this information, we can then use modelling to predict the cell behaviour and to automatically control the bioreactor conditions to improve production.As a demonstration, we will focus on monitoring the buildup of lactic acid which is a byproduct of anaerobic (oxygen-poor) reaction conditions; excess lactic acid is toxic, and can limit the performance of (or even kill) the cells that produce it. By engineering the cells to glow based on how much lactic acid there is nearby, we can monitor the reaction and either increase the amount of oxygen added to the reaction, stir the tank, or even redesign the reactor itself to avoid local differences in the reaction conditions. The case of lactic acid is just an example; future cells might report changes in temperature, shear stress, oxygenation or any other parameter, with a different-coloured glow for each.Overall, this project represents the first step towards a new frontier where the cells in a bioreactor not only produce the chemicals we want, but tell us what is going wrong in the reaction and how to fix it. The result is a reactor that can make complex chemicals much more cheaply, and given how much of our modern world relies on these chemicals, that can have subtle but pronounced benefits throughout the global economy.