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生命科学中大多数研究项目的基石。人们越来越认识到，对细胞功能的准确理解需要对组织中的原代细胞进行研究，而不能用培养或其他体外系统中转化的细胞系来准确建模。然而，在现有的生物样本中，大多数原代细胞的数量都很少，而且本质上是异质性的。对于适应性免疫系统的细胞、B淋巴细胞和T淋巴细胞来说尤其如此。这些细胞中的每一个在发育过程中都获得了一个独特的结构和特异性的抗原识别受体，并在参与免疫反应的过程中经历了进一步的分化、极化和功能规范。精确的受体刺激阈值决定了细胞的命运，影响淋巴细胞如何参与免疫反应或强制细胞无能和凋亡。传统的实验方法，如Western blotting，分析的是大量的细胞群体，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的分子分析对于准确了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。*这项应用汇集了来自麦吉尔大学的一支杰出的免疫学家团队，他们研究免疫反应中调节B和T细胞功能的分子机制，并面临着小而异生物样本中蛋白质定量的共同挑战。应这一应用的要求，Milo Single-Cell Western系统是市场上唯一允许在单细胞分辨率下对小生物样本进行蛋白质定量的同类仪器，目前麦吉尔大学和蒙特利尔地区尚不提供。因此，迫切需要该设备，并有可能从根本上推进申请者的研究计划，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。这种洞察力是现有或替代设备和方法无法获得的。重要的是，申请者在HQP培训方面有着良好的记录，目前他们的联合研究团队包括30多名受训人员。这些受训人员中的大多数都在从事需要对小细胞和异质细胞样本中的蛋白质进行量化的项目，因此将直接和强烈地受益于所要求的Milo Single-Cell Western系统。HQP的好处将包括直接获得新的技术技能，提高研究生产率，以及他们的工作对研究领域的更大影响。*总体而言，所需设备为学术领域和HQP培训带来了重大好处。**

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.**