使用快速有效的脑损伤生物标志物分析，改善创伤治疗结果
"“我们很幸运能够在研究工作的早期试用这些 Luminex 猪类样本分析组。当我们看到检测板的测量结果，就知道我们找到了一直在寻找的分析组。这些组合最终对研究结果产生了巨大影响。”"
- Elizabeth Rick, 俄勒冈健康与科学大学 (OHSU) 研究科学家
LUMINEX® 分析帮助推进脑损伤生物标志物分析研究
在美国，创伤性脑损伤 (TBI) 是造成死亡和残疾的主要原因之一，每年约有 280 万人因为 TBI 就医。1TBI 的可用治疗选择有限，当前策略主要侧重于针对初始症状和继发性影响进行反应性管理。
Elizabeth Rick 是俄勒冈健康与科学大学 (OHSU) 的研究科学家，在外科学创伤、重症监护和急症护理部门工作。OHSU 创伤部门正在开展一项临床和基础研究计划，专注于研发创新的治疗和护理方法，造福 OHSU 及世界各地的创伤患者。
最近的研究（包括一项由 OHSU 创伤实验室指导、为头部创伤患者使用 TXA 的大型随机化临床研究）表明，早期使用氨甲环酸 (TXA) 可降低死亡率并改善神经结局。TXA 是一种强效抗纤溶药，可以抑制血块被酶溶解。2研究团队的核心重点是深入了解 TXA 与 TBI 患者内皮受损之间的关系。
更高的分析灵敏度助力发现 TBI 生物标志物
死亡率的降低可以归因于颅内出血 (ICH) 减少，研究人员越发关注 ICH 减少的机制和 TXA 在帮助预防继发性损伤方面的作用。3OHSU 研究团队在最近的一项研究中，考察了早期 TXA 给药对内皮单层组织病变循环标志物的影响；对中重度创伤性脑损伤 (MS-TBI) 和 ICH 患者的许多内皮标志物和细胞因子浓度进行了分类记录。
快速有效地定量检测生物标志物并将其与特定结局相关联，这是所有转化研究领域的一项关键要求。在这种情况下，Bio-Techne 的 Luminex Discovery 分析为 OHSU 团队带来了理想的解决方案。
多重分析的筛选能力不仅节约成本，而且让研究人员能够快速有效地同时分析内皮和凝血活化情况——这是保持内皮单层组织完整性的两个关键因素。同时，Luminex 灵活的配置选项让研究人员可以从大量的分析类型选项中进行选择，以构建其定制分析组。
“Luminex 分析组卓越的准确度和灵敏度发挥了重要作用，让我们获得了所需的高质量数据。”Rick 表示，“当我们发现了目标并看到了测出的浓度，这增加了我们对结果的信心——即内皮单层组织功能得到改善。”
找到正确的猪类样本分析
创伤研究组还使用猪类模型来深入研究创伤后内皮单层组织病变和内皮损伤。这一研究重点需要稳健的分析，特别是在动物模型中，但是合适的猪类样本分析组通常很难找到，因为猪类模型不像啮齿动物模型那么常用。
“猪类模型不是通用的模型，所以没有很多的多重分析选择。”Rick 讲道，“我们与几个供应商合作，尝试以多重分析形式进行猪细胞因子分析，但都失败了。我们进行了分析，但没有获得一致的可用浓度。”
最近，创伤研究团队开展了一项研究，分析干细胞治疗在预防或降低创伤性出血和损伤后所引发的急性呼吸窘迫综合征 (ARDS) 的效果，再次发现他们需要合适的猪类样本分析组来开展多重分析，满足检测要求。由于与 Bio-Techne 的密切合作关系，他们最终找到了一直在寻找的分析方法。
2019 年，她与 Bio-Techne 销售代表交谈时，该代表碰巧提到了一些正在开发中的新型 Luminex 猪类样本分析组。Rick 灵机一动，问销售代表是否可以提前使用仍在进行 β 测试的一些检测板。
“我们很幸运能够在研究工作的早期试用这些 Luminex Discovery 猪类样本分析。”Rick 表示，“当我们看到检测板的测量结果，就知道我们找到了一直在寻找的分析组。这些组合最终对研究结果产生了巨大影响。”
在早期建立对研究结果的信心
Rick 针对有脑损伤、内皮单层组织病变和创伤性出血影响的生物标志物开展了广泛研究，对所采集的样本进行分析，使用过各种不同的分析方法和平台。在最近的一个项目中，Rick 和她的团队使用了 Bio-Techne 的 Luminex 分析和 Quantikine™ ELISA 的组合。
对于这一项目，Rick 的任务是培训当地大学的一群实习生设置和进行多个酶联免疫吸附试验 (ELISA)。Rick 说，尽管这些分析的设置和操作对他们来说都是全新的，但没关系，因为这些分析的操作流程简单，准确度出众。
“我们设置并运行了 10 个检测板。”Rick 介绍，“曲线很漂亮，每次的结果都惊人的准确。与大学生一起工作很有趣，他们现在以为所有 ELISA 都能这样完美进行。”
在准确度和一致性方面获得优势
事实证明，Luminex 和 ELISA 分析简明易懂，为研究团队带来重要优势。
“从我们第一次看到 Bio-Techne 检测板开始，我们就发现操作流程和程序的清晰度明显不同。”Rick 评价。“在总体分析的一致性和准确度方面，没有可以与它相比的。”Rick 赞叹，“我们一直使用 Bio-Techne 的产品，因为我们在别处都找不到这么高质量的分析。”
从基于酶标板的 ELISA 转为 Luminex Discovery 猪类样本分析，使 Rick 及其团队能够充分利用多重分析的效率和通量。
“不仅是酶标板的成本，还有设置更多 ELISA 操作所需的时间。”Rick 讲道，“我们内部员工人手有限，所以当我们找到 Luminex 猪类样本分析组的时候，真的很兴奋。”
能够在一次分析中评估多种分析物，这让 Rick 及其团队能够审慎使用宝贵的样本材料，获得更多有用信息，而无需根据样本量来安排分析的优先顺序，并且这些样本还可以用于未来研究。
有了正确的分析工具，Rick 可以继续致力于造福患者的转化研究，同时她可以验证通过各种研究和不同分析平台得到的结果。未来，研究团队计划使用 Luminex Discovery 分析人类模型样本，针对 TBI 和创伤患者的内皮标志物和细胞因子开展更多分析。
参考文献
- Taylor CA, Bell JM, Breiding MJ, Xu L. (2007 and 2013) Traumatic brain injury-related emergency department visits, hospitalizations, and deaths—United States,MMWR Surveill Summ. 66(9):1–16.
- Zehtabchi S, Abdel Baki SG, Falzon L, Nishijima DK. (2014) Tranexamic acid for traumatic brain injury: a systematic review and meta-analysis. Am J Emerg Med. 32(12):1503–1509.
- Jokar A, Ahmadi K, Salehi T, Sharif-Alhoseini M, Rahimi-Movaghar V. (2017) The effect of tranexamic acid in traumatic brain injury: a randomized controlled trial. Chin J Traumatol. 20(1):49–51.
- Akwii RG, Sajib MS, Zahra FT, Mikelis CM. (2019) Role of angiopoietin-2 in vascular physiology and pathophysiology. Cells. 8(5):471.
Improving Discovery Of TBI Biomarkers With Better Assay Sensitivity
While decreased mortality has been attributed to reduction in intracranial hemorrhage (ICH), there’s growing interest in characterizing the possible mechanisms associated with this reduction and the effect of TXA in helping to prevent secondary injury.3 In a recent study examining the effect of early TXA administration on circulating markers of endotheliopathy, the research team at OHSU was able to catalog many of the endothelial markers and cytokine concentrations in patients with moderate-to-severe traumatic brain injury (MS-TBI) and ICH.
The need to quantify biomarkers quickly and efficiently and correlate them to specific outcomes is a critical requirement across all areas of translational research. In this case, the OHSU team found the optimum solution with Luminex Discovery assays from Bio-Techne.
The cost-efficient screening capabilities of the multiplex format provided researchers with a fast and effective way to simultaneously profile endothelial and coagulation activation—two key factors in the preservation of endothelial integrity. Meanwhile, the flexible Luminex configuration options allowed researchers to choose from a large menu of analytes to build their custom assay panel.
“The superior accuracy and sensitivity of the Luminex panels proved instrumental in giving us the high-quality data we needed,” Rick said. “Once we identified our targets and could see the concentrations we were getting, this increased our confidence in the results indicating improved endothelial function.”
Finding The Right Porcine Assay
The Trauma Research group additionally uses a pig model to improve insight into endotheliopathy and the endothelial damage following trauma. This focus demands robust assays, particularly in the animal model but suitable porcine panels have been difficult to find largely because the porcine model is not as frequently used as the rodent models.
“The porcine model is not a universal model so there are not a lot of multiplexing options,” Rick said.
“We’ve worked with several vendors to try and set up a porcine cytokine assay in a multiplex format, but all of them have come up short. We would run the assay and there would be no consistent, robust concentrations on the plate.”
In one recent study analyzing the effect of stem cell treatment to prevent or lessen the possibility of acute respiratory distress syndrome (ARDS) following traumatic hemorrhage and injury, the Trauma Research team found themselves once again searching for a suitable porcine panel in a multiplex assay format that could meet their detection requirements. They ultimately found what they were looking for, thanks to its close partnership with Bio-Techne.
While talking with her Bio-Techne sales rep in 2019, the rep happened to mention some new Luminex porcine panels that were currently in development. Thinking quickly, Rick asked the rep if it was possible to get early access to some of the plates that were still in beta testing.
“We were fortunate to be able to test these Porcine Luminex Discovery Assays early in our research effort,” Rick said. “Once we could see the measurements we were getting on the plates, we knew we had found what we were looking for. The panels ended up making a huge difference in the outcome of the study.”
Building Early Confidence In Their Research Results
As part of her comprehensive research analyzing collected samples for biomarkers of brain injury, endotheliopathy, and traumatic hemorrhage effects, Rick has experimented with a variety of different assays and platforms. In one recent project, Rick and her team employed a combination of Luminex assays and Quantikine™ ELISAs from Bio-Techne.
For this particular project, Rick was tasked with training a group of interns from the local university on how to set up and run several of the ELISAs.
Even though the setup and operation of the assays were all new to them, it didn't matter, Rick said, because of the easy-to-follow protocols and the exceptional accuracy of the assays. “We set up and ran 10 plates,” Rick said. “The curves were beautiful, and the results were unbelievably accurate every time. It was really fun to work with this group —they now think that all ELISAs run that flawlessly.”
Gaining An Accuracy And Consistency Advantage
The ease of use and transparency of both the Luminex and ELISA assays has proven to be an important advantage for the research team.
“From the first time we looked at the Bio-Techne plates, we noticed a clear difference in the clarity of the protocols and procedures,” Rick said. “When it comes to overall assay consistency and accuracy, there’s no comparison.” “We keep coming back to Bio-Techne because we can’t find that quality anywhere else.”
Transitioning from plate-based ELISAs to the Porcine Luminex Discovery assays allowed Rick and her team to take advantage of the efficiency and throughput advantages of the multiplex format.
“It’s not just the cost of the plates, but also the time required to set up a larger number of ELISAs,” Rick said. “We don’t have a large in-house staff, so when we found the Luminex porcine panels, it was really exciting.”
Assessing multiple analytes in a single assay also allows Rick and her team to be judicious with materials while generating much more useful information, alleviating the need to prioritize assays based on the volume of precious samples and allowing for future studies using those same samples.
With access to the right analytical tools, Rick can continue doing translational research that impacts lives while remaining confident in her ability to validate results across different types of studies and between different assay platforms. In the future, the research team plans to use the Luminex Discovery assays with human models to perform additional analysis on endothelial markers and cytokines in TBI and trauma patients.
REFERENCES
- Taylor CA, Bell JM, Breiding MJ, Xu L. (2007 and 2013) Traumatic brain injury-related emergency department visits, hospitalizations, and deaths—United States, MMWR Surveill Summ. 66(9):1–16.
- Zehtabchi S, Abdel Baki SG, Falzon L, Nishijima DK. (2014) Tranexamic acid for traumatic brain injury: a systematic review and meta-analysis. Am J Emerg Med. 32(12):1503–1509.
- Jokar A, Ahmadi K, Salehi T, Sharif-Alhoseini M, Rahimi-Movaghar V. (2017) The effect of tranexamic acid in traumatic brain injury: a randomized controlled trial. Chin J Traumatol. 20(1):49–51.
- Akwii RG, Sajib MS, Zahra FT, Mikelis CM. (2019) Role of angiopoietin-2 in vascular physiology and pathophysiology. Cells. 8(5):471.