Chaturvedi Shalini, Siegel Derick, Wagner Carrie L, Park Jaehong, van de Velde Helgi, Vermeulen Jessica, Fung Man-Cheong, Reddy Manjula, Hall Brett, Sasser Kate
Oncology, Translational Research, Janssen Pharmaceutical Companies of Johnson & Johnson, Spring House, Pennsylvania.
Oncology, Discovery, Janssen Pharmaceutical Companies of Johnson & Johnson, Spring House, Pennsylvania.
Br J Clin Pharmacol. 2015 Oct;80(4):687-97. doi: 10.1111/bcp.12652. Epub 2015 Jun 4.
Interleukin-6 (IL-6), a multifunctional cytokine, exists in several forms ranging from a low molecular weight (MW 20-30 kDa) non-complexed form to high MW (200-450 kDa), complexes. Accurate baseline IL-6 assessment is pivotal to understand clinical responses to IL-6-targeted treatments. Existing assays measure only the low MW, non-complexed IL-6 form. The present work aimed to develop a validated assay to measure accurately total IL-6 (complexed and non-complexed) in serum or plasma as matrix in a high throughput and easily standardized format for clinical testing.
Commercial capture and detection antibodies were screened against humanized IL-6 and evaluated in an enzyme-linked immunosorbent assay format. The best antibody combinations were screened to identify an antibody pair that gave minimum background and maximum recovery of IL-6 in the presence of 100% serum matrix. A plate-based total IL-6 assay was developed and transferred to the Meso Scale Discovery (MSD) platform for large scale clinical testing.
The top-performing antibody pair from 36 capture and four detection candidates was validated on the MSD platform. The lower limit of quantification in human serum samples (n = 6) was 9.77 pg l(-1) , recovery ranged from 93.13-113.27%, the overall pooled coefficients of variation were 20.12% (inter-assay) and 8.67% (intra-assay). High MW forms of IL-6, in size fractionated serum samples from myelodysplastic syndrome and rheumatoid arthritis patients, were detected by the assay but not by a commercial kit.
This novel panoptic (sees all forms) IL-6 MSD assay that measures both high and low MW forms may have clinical utility.
白细胞介素-6（IL-6）是一种多功能细胞因子，存在多种形式，从低分子量（分子量20 - 30 kDa）的非复合形式到高分子量（200 - 450 kDa）的复合物形式。准确的基线IL-6评估对于理解针对IL-6的治疗的临床反应至关重要。现有检测方法仅测量低分子量的非复合IL-6形式。本研究旨在开发一种经过验证的检测方法，以高通量且易于标准化的临床检测形式，准确测量血清或血浆基质中的总IL-6（复合和非复合形式）。
针对人源化IL-6筛选商业捕获和检测抗体，并以酶联免疫吸附测定形式进行评估。筛选最佳抗体组合，以鉴定在存在100%血清基质的情况下背景最低且IL-6回收率最高的抗体对。开发了基于板的总IL-6检测方法，并转移至Meso Scale Discovery（MSD）平台进行大规模临床检测。
在MSD平台上验证了从36种捕获抗体和4种检测候选抗体中表现最佳的抗体对。人血清样本（n = 6）中的定量下限为9.77 pg l(-1)，回收率在93.13 - 113.27%之间，总体合并变异系数为20.12%（批间）和8.67%（批内）。该检测方法可检测骨髓增生异常综合征和类风湿关节炎患者的大小分级血清样本中的高分子量IL-6形式，但商业试剂盒无法检测。
这种能够检测高分子量和低分子量形式的新型全景式（能检测所有形式）IL-6 MSD检测方法可能具有临床应用价值。
