We propose to establish a new Amnis ImageStreamX Core. Microscopy can image cytological rearrangements, cell:cell interactions, protein and membrane localization, and cell signaling events. However, such methods are neither high throughput nor quantitative. Nor are they adapted for real-time signaling or facile for the separation of defined cell types from mixtures. With these limitations in mind, Yale School of Medicine (YSM) acquired through a shared instrument grant an Amnis ImageStreamX. The Amnis ImageStreamX is a multispectral imaging instrument that acquires 12 channels of cellular imagery. By collecting large numbers of images per sample and providing numerical representation of image-based features, the ImageStreamX combines the per cell information content of standard microscopy with the statistical significance afforded by large sample sizes in standard flow cytometry. The best applications for the ImageStreamX take advantage of its abilities to locate and quantitate the distribution of signals on, within or between cells. This is the only such instrument at YSM. While there have been important discoveries made with the Amnis, the instrument is clearly underutilized. This is despite an enthusiastic initial response during a trial period wherein an expert from Amnis assisted in planning experiments, running samples and analyzing data on-site for 3 days. During this short time, preliminary data were obtained for 10 projects from 10 independent investigators. Whereas this early success was achieved with an expert on-site operator, the model applied for ongoing use of the Amnis was to treat as a user-operated 4-colorfiow cytometer. This model was not self-sustaining. Key aspects of the technology and in particular the required analysis of massive data sets rendered optimal use of the Amnis more complicated than a flow cytometer and therefore less accessible. This proposal seeks to remedy this by applying lessons learned by the P30 2P Core for which a dedicated and experienced operator participates in all phases of the work including experimental planning, training of users, imaging, assistance and training in data analysis. We propose to apply the same model to the Amnis so as to augment its impact on the study of rheumatic and immunological diseases, and the pathways that mediate their pathogenesis.
Autoimmunity and rheumatologic diseases are a major cause of illness, disability and even death. Tools that enable research into these diseases are essential for progress in understanding disease etiology and in developing new therapies. The Amnes ImageStreamX Core is such a tool and this proposal seeks to make it more available to investigators studying autoimmune and rheumatic diseases.
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