PDF(Pubmed)
Abstract：
Several CD19 targeted antibody-based therapeutics are currently available for patients with diffuse large B-cell lymphoma (DLBCL), including the Fc-modified antibody immunotherapy tafasitamab. This therapeutic landscape warrants the evaluation of potential sequencing approaches. Prior to a subsequent CD19-targeted therapy, CD19 expression on tafasitamab-treated patient biopsy samples may be assessed. However, no standardized methods for its detection are currently available. In this context, selecting a tafasitamab-competing CD19 detection antibody for immunohistochemistry (IHC) or flow cytometry (FC) may lead to misinterpreting epitope masking by tafasitamab as antigen loss or downregulation.
We analyzed a comprehensive panel of commercially available CD19 detection antibody clones for IHC and FC using competition assays on tafasitamab pre-treated cell lines. To remove bound tafasitamab from the cell surface, an acidic dissociation protocol was used. Antibody affinities for CD19 were measured using Surface Plasmon Resonance (SPR) or Bio-Layer Interferometry (BLI).
While CD19 was successfully detected on tafasitamab pre-treated samples using all 7 tested IHC antibody clones, all 8 tested FC antibody clones were confirmed to compete with tafasitamab. An acidic dissociation was demonstrated essential to circumvent CD19 masking by tafasitamab and avoid false negative FC results.
The current study highlights the importance of selecting appropriate CD19 detection tools and techniques for correct interpretation of CD19 expression. The findings presented herein can serve as a guideline to investigators and may help navigate treatment strategies in the clinical setting.
We analyzed a comprehensive panel of commercially available CD19 detection antibody clones for IHC and FC using competition assays on tafasitamab pre-treated cell lines. To remove bound tafasitamab from the cell surface, an acidic dissociation protocol was used. Antibody affinities for CD19 were measured using Surface Plasmon Resonance (SPR) or Bio-Layer Interferometry (BLI).
While CD19 was successfully detected on tafasitamab pre-treated samples using all 7 tested IHC antibody clones, all 8 tested FC antibody clones were confirmed to compete with tafasitamab. An acidic dissociation was demonstrated essential to circumvent CD19 masking by tafasitamab and avoid false negative FC results.
The current study highlights the importance of selecting appropriate CD19 detection tools and techniques for correct interpretation of CD19 expression. The findings presented herein can serve as a guideline to investigators and may help navigate treatment strategies in the clinical setting.
摘要：
■目前有几种基于CD19靶向抗体的疗法可用于弥漫性大B细胞淋巴瘤（DLBCL）患者，包括Fc修饰的抗体免疫疗法tafasitamab。这种治疗前景需要评估潜在的测序方法。在随后的CD19靶向治疗之前，可以评估他法他单抗治疗的患者活检样品上的CD19表达。然而,目前尚无标准化的检测方法。在这种情况下，选择他法他单抗竞争性CD19检测抗体进行免疫组织化学(IHC)或流式细胞术(FC),可能会导致他法他单抗将表位掩蔽误解为抗原丢失或下调.
我们使用在他法他单抗预处理的细胞系上的竞争测定分析了用于IHC和FC的市售CD19检测抗体克隆的综合组。要从细胞表面去除结合的他法他单抗，使用酸性解离方案。使用表面等离子体共振(SPR)或生物层干涉法(BLI)测量对CD19的抗体亲和力。
■虽然使用所有7个测试的IHC抗体克隆成功检测到他法他单抗预处理样品上的CD19，所有8个测试的FC抗体克隆均被证实与他法他单抗竞争.证明酸性解离对于规避tafasitamab的CD19掩蔽和避免FC假阴性结果是必不可少的。
■当前的研究强调了选择适当的CD19检测工具和技术以正确解释CD19表达的重要性。本文提出的发现可以作为研究者的指南，并且可以帮助在临床环境中导航治疗策略。
我们使用在他法他单抗预处理的细胞系上的竞争测定分析了用于IHC和FC的市售CD19检测抗体克隆的综合组。要从细胞表面去除结合的他法他单抗，使用酸性解离方案。使用表面等离子体共振(SPR)或生物层干涉法(BLI)测量对CD19的抗体亲和力。
■虽然使用所有7个测试的IHC抗体克隆成功检测到他法他单抗预处理样品上的CD19，所有8个测试的FC抗体克隆均被证实与他法他单抗竞争.证明酸性解离对于规避tafasitamab的CD19掩蔽和避免FC假阴性结果是必不可少的。
■当前的研究强调了选择适当的CD19检测工具和技术以正确解释CD19表达的重要性。本文提出的发现可以作为研究者的指南，并且可以帮助在临床环境中导航治疗策略。
