Sandwich ELISA is an ideal antigen detection and quantification assay. Recently, it was used as the basic concept for high technology diagnostics. The specificity of the assay depends on the exclusion of detection cross-reactivity which arises from using two antibodies developed in different species. Since mice and rats are the common laboratory animals used to develop antigen specific antibodies. Therefore, the questions we addressed here were (1) can one use antigen-specific antibodies raised in mice and rats in the same assay to specifically detect/quantify antigens? and (2) which antibodies of the two rodents should be placed for capturing and for detection in the antigen-detection sandwich? Direct ELISA assay was used to assess for the specific reaction of the HRP-conjugated antibody to the target serum. First reaction was to compare between either conjugate anti-rat IgG (homologous) or anti-mouse IgG (heterologous) for the detection of rat sera IgG. Following the dilution factor optimization, the O.D. were 0.744±0.051 and 0.604±0.05, respectively (p= .004). The difference in mean O.D. of 0.14 reflected an unaccepted non-specific reaction. The second reaction was to compare between either conjugate anti-mouse IgG (homologous) and anti-rat IgG (heterologous) for the detection of mouse sera IgG. The recorded O.D. were 0.9414±0.14 and 0.317 ±0.141, respectively (p= .0002). The improved difference in mean O.D. of 0.624 reflecting a minimized cross-reaction. Our results suggest that it is possible to use both Swiss albino mice and albino rats in a single sandwich ELISA, given that the captured antibody species to be from the Swiss albino mice and the detection antibody to be from the albino rat. The described working details are limited to the source of the antibodies used in the study. However, the approach stresses on the importance of such optimization steps before making any interpretations based on the antigen detection. To our knowledge, this study is the first to cover the optimal order of the capturing and the detection antibodies in a sandwich ELISA assay. In addition to addressing the possible interfering cross-reactivity that result from using mouse and rat serum antibodies in a single assay.

中文翻译：

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小鼠和大鼠免疫球蛋白 G 之间的交叉反应：它在夹心 ELISA 中重要吗？

夹心 ELISA 是一种理想的抗原检测和定量分析。最近，它被用作高科技诊断的基本概念。检测的特异性取决于排除因使用在不同物种中开发的两种抗体而产生的检测交叉反应。由于小鼠和大鼠是用于开发抗原特异性抗体的常见实验室动物。因此，我们在这里解决的问题是 (1) 是否可以使用在同一测定中在小鼠和大鼠中产生的抗原特异性抗体来特异性检测/量化抗原？(2) 在抗原检测夹心中应该放置两种啮齿动物的哪些抗体进行捕获和检测？直接 ELISA 测定用于评估 HRP 偶联抗体与目标血清的特异性反应。第一个反应是比较用于检测大鼠血清 IgG 的缀合物抗大鼠 IgG（同源）或抗小鼠 IgG（异源）。稀释因子优化后，OD 分别为 0.744±0.051 和 0.604±0.05 (p = .004)。0.14 的平均 OD 差异反映了不可接受的非特异性反应。第二个反应是比较用于检测小鼠血清 IgG 的偶联抗小鼠 IgG（同源）和抗大鼠 IgG（异源）。记录的 OD 分别为 0.9414±0.14 和 0.317±0.141 (p = .0002)。平均 OD 的改进差异为 0.624，反映了最小化的交叉反应。我们的结果表明，可以在单个夹心 ELISA 中同时使用瑞士白化小鼠和白化大鼠，鉴于捕获的抗体种类来自瑞士白化小鼠，检测抗体来自白化大鼠。所描述的工作细节仅限于研究中使用的抗体来源。然而，在基于抗原检测做出任何解释之前，该方法强调了此类优化步骤的重要性。据我们所知，这项研究是第一个涵盖夹心 ELISA 测定中捕获和检测抗体的最佳顺序的研究。除了解决在单一检测中使用小鼠和大鼠血清抗体可能导致的干扰性交叉反应之外。在根据抗原检测做出任何解释之前，该方法强调了此类优化步骤的重要性。据我们所知，这项研究是第一个涵盖夹心 ELISA 测定中捕获和检测抗体的最佳顺序的研究。除了解决在单一检测中使用小鼠和大鼠血清抗体可能导致的干扰性交叉反应之外。在根据抗原检测做出任何解释之前，该方法强调了此类优化步骤的重要性。据我们所知，这项研究是第一个涵盖夹心 ELISA 测定中捕获和检测抗体的最佳顺序的研究。除了解决在单一检测中使用小鼠和大鼠血清抗体可能导致的干扰性交叉反应之外。