Molecular Analysis of the Diversity of Lymphoid Cell Types and Functions at Single-Cell Resolution: ProteinSimple Single-Cell Western System.

细胞的蛋白质组成决定了它的功能特性。因此，细胞蛋白质的定量测量是大多数生命科学研究项目的基石。人们越来越认识到，要准确地了解细胞功能，需要对来自组织的原代细胞进行研究，而不能用培养或其他体外系统中的转化细胞系精确地建模。然而，大多数原代细胞在现有的生物样品中数量非常少，并且本质上是异质的。对于适应性免疫系统细胞，B淋巴细胞和T淋巴细胞尤其如此。这些细胞在发育过程中获得具有独特结构和特异性的抗原识别受体，并在参与免疫应答过程中进一步分化、极化和功能规范。精确的受体刺激阈值决定细胞命运，影响淋巴细胞如何参与免疫反应或加强细胞能量和凋亡。传统的实验方法，如Western blotting，分析的是大量细胞群，不能准确地捕捉适应性免疫系统细胞的功能复杂性。因此，在单细胞分辨率下对稀有细胞类型进行分子分析对于准确理解淋巴细胞功能以及适应性免疫反应和免疫调节机制至关重要。******该应用程序汇集了来自麦吉尔大学的杰出免疫学家团队，研究免疫应答中调节B细胞和T细胞功能的分子机制，并面临小样本和异质生物样品中蛋白质定量的共同挑战。该申请要求的Milo单细胞Western系统是市场上唯一一种允许在单细胞分辨率下对小生物样品进行蛋白质定量的仪器，目前在麦吉尔大学和蒙特利尔地区还没有。因此，该设备是迫切需要的，并且有可能从根本上推进申请人的研究计划，为B和T淋巴细胞发育和适应性免疫反应的分子机制提供新的见解。现有的或可替代的设备和方法无法获得这种洞察力。******重要的是，申请人在HQP培训方面有着出色的记录，目前他们的研究团队包括30多名学员。这些受训人员中的大多数正在从事需要在小细胞和异质细胞样品中进行蛋白质定量的项目，因此将直接和强烈地受益于所要求的Milo单细胞Western系统。对HQPs的好处将包括直接获得新的技术技能，提高研究生产力，并增加其工作对研究领域的影响。******总体而言，所要求的设备为学术领域和HQP培训提供了主要好处。**

Protein composition of a cell defines its functional properties. Quantitative measurements of cellular proteins are therefore the cornerstone of most research programs in life sciences. It is increasingly recognized that an accurate understanding of cellular function requires studies with primary cells from the tissues, and can not be accurately modeled with transformed cell-lines in culture or other in vitro systems. However, most primary cells are found in very low numbers in the available biological samples, and are heterogeneous in nature. This is especially true for the cells of adaptive immune system, B and T lymphocytes. Each of these cells acquires an antigen recognition receptor of a unique structure and specificity during its development, and undergoes further differentiation, polarization, and functional specification during its engagement in immune response. Precise receptor stimulation thresholds dictate cell-fate, affecting how a lymphocyte engages in immune response or enforcing cell-anergy and apoptosis. The traditional experimental methods, like Western blotting, analyze bulk cell populations and do not accurately capture the functional complexity of the cells of adaptive immune system. The molecular analysis of rare cell types at single cell resolution is therefore critical for accurate understanding of lymphocyte functions and the mechanisms of adaptive immune response and immune regulation.******This application brings together a team of outstanding immunologists from across McGill University, studying the molecular mechanisms that regulate B and T cell functions in immune response, and facing the common challenge of protein quantification in small and heterogeneous biological samples.The Milo Single-Cell Western System, requested on this application, is the only instrument of its kind on the market allowing protein quantification in small biological samples at single cell resolution, and is not currently available at McGill University and the Montreal area. The equipment is therefore urgently needed and has the potential to fundamentally advance the research programs of the applicants, providing novel insight into the molecular mechanisms of B and T lymphocyte development and adaptive immune response. Such insight can not be obtained with existing or alternative equipment and methodologies.******Importantly, the applicants have an excellent track-record in HQP training, and currently their combined research teams include over 30 trainees. Most of these trainees are working on projects that require protein quantification in small and heterogeneous cell samples, and will therefore directly and strongly benefit from the requested Milo Single-Cell Western System. The benefits to the HQPs will include the direct acquisition of new technical skills, enhanced research productivity, and increased impact of their work on the research field. ******Overall, the requested equipment offers major benefits to the academic field and HQP training.**