血清游离轻链 (FLC) 是诊断和监测浆细胞肿瘤患者的重要临床生物标志物。目前广泛使用的免疫分析方法对总血清FLC进行定量，其中包括单克隆FLC以及多克隆背景中的FLC。患有慢性疾病、炎症性疾病或肾功能障碍的患者的总 FLC 可能会升高，从而导致结果不明确。这些患者可能会受益于单克隆 FLC 的直接测量。本研究的目的是开发一种将探针萃取 (OPEX) 与液相色谱-高分辨率质谱 (LC-HR-MS)（简称 OPEX-MS）相结合的方法，以直接确定 FLC 的克隆性。 OPEX 免疫捕获使用负载有抗 kappa 或抗 lambda 轻链抗体的微探针进行。捕获的蛋白质通过反相 LC 进行分离，并通过 HR-MS 进行分析。根据免疫分析 FLC 结果对来自独特患者的四组样本进行了测试。 OPEX-MS 方法中的 LC-HR-MS 分析提供了每个克隆的独特保留时间以及 FLC 单体和二聚体的解卷积质量。研究发现，49 个 kappa FLC 升高的样本中，有 16 个 (33%) 以及 100 个 kappa 和 lambda FLC 双升高样本中的 83 个 (83%) 没有单克隆 FLC，这与通常存在单克隆 FLC 的知识是一致的。 FLC 免疫测定结果轻度升高的样本中没有克隆群体。OPEX-MS 方法可以作为一种补充方法，直接确定 FLC 免疫测定结果难以解释的患者的克隆性。© 诊断协会

Serum free light chains (FLCs) are an essential clinical biomarker for the diagnosis and monitoring of patients with plasma cell neoplasms. The current widely used immunoassay methods quantify total serum FLCs, which include monoclonal FLCs as well as FLCs in the polyclonal background. Patients with chronic diseases, inflammatory disorders, or renal dysfunction can have elevated total FLCs that lead to ambiguous results. These patients may benefit from a direct measurement of monoclonal FLCs. The purpose of this study was to develop a method that couples on-probe extraction (OPEX) with liquid chromatography-high-resolution mass spectrometry (LC-HR-MS), abbreviated to OPEX-MS, to directly determine the clonality of FLCs.OPEX immunocapture was performed using microprobes loaded with anti-kappa or anti-lambda light chain antibodies. Captured proteins were separated by reversed-phase LC and analyzed by HR-MS.Four cohorts of samples from unique patients were tested based on immunoassay FLC results. The LC-HR-MS analysis in the OPEX-MS method provides both a unique retention time along with deconvoluted masses of FLC monomers and dimers for each clone. The study found that 16 out of 49 (33%) kappa FLC elevated samples as well as 83 out of 100 (83%) dual kappa and lambda FLC elevated samples did not have monoclonal FLCs, which is consistent with the knowledge that there is often no clonal population in samples with mildly elevated FLC immunoassay results.The OPEX-MS method can serve as a complementary approach to directly determine clonality in patients with difficult-to-interpret FLC immunoassay results.© Association for Diagnostics & Laboratory Medicine 2024. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.