金抗体替代ELISA中的天然抗体用于溶菌酶检测

Goldbody as a Replacement of Natural Antibody in Enzyme Linked Immunosorbent Assay for Detection of Lysozyme

目的酶联免疫吸附测定(ELISA)被广泛用于抗体或抗原的检测,并被视为临床实践中的金标准,可提供相对可靠、灵敏和特异的检测结果。ELISA的本质是抗原与相应抗体之间的特异性相互作用。然而,天然抗体固有的不稳定性是ELISA的一个难以克服的弱点,并可能导致检测结果的重现性差甚至错误的诊断结果。本课题组先前应用构象工程方法开发了一种基于金纳米粒子的人工抗体(简称金抗体)。金抗体可以像天然抗体一样特异性地与抗原相互作用,并且具备远优于天然抗体的稳定性。出色的稳定性使金抗体可能成为天然抗体更好的替代物,用于ELISA中。方法经过必要的优化并与辣根过氧化物酶(HRP)耦联后,制得酶标金抗体10HRP-(Au-400P1),然后用酶标金抗体代替天然酶标抗体用于ELISA检测中。结果通过一系列的实验证明,抗溶菌酶金抗体可用于ELISA特异性检测1~16 mg/L范围内的鸡蛋清溶菌酶(HEWL)样品。结论金抗体可以替代天然抗体用于ELISA检测,并具有优于传统ELISA法的检测准确性和一致性。

Objective Enzyme linked immunosorbent assay(ELISA)has been widely used for detection of antibodies or antigens,and is regarded as the gold standard in clinical diagnosis,which can provide relatively reliable,sensitive and specific results.The essence of ELISA is the specific interaction between antigens and the corresponding antibodies.However,the inherent instability of natural antibodies is the Achilles heel of ELISA,which may lead to poor reproducibility or even false results.Previously,our group created a nova gold nanoparticle-based artificial antibody,denoted as goldbody.Goldbody can specifically interact with antigens like natural antibodies,but has much better stability than that of natural antibodies.The excellent stability of goldbody makes it a potential replacement of natural antibodies in ELISA assays.Methods Herein,we demonstrate with a series of experiments that goldbody is indeed a good replacement of natural antibodies in ELISA for antigen detection.Results After necessary optimization and conjugation of horseradish peroxidase(HRP),the 13 nm HRP-labeled anti-hen egg white lysozyme(HEWL)goldbody can be used in ELISA assay to detect HEWL in the range of 1-16 mg/L.Conclusion Goldbodies can indeed replace natural antibodies in ELISA for a more accurate and consistent detection of antigens.