The aim of this work was to investigate the complex phenomena underlying the adsorption of an anti-human IL-8 (anti-IL8) monoclonal antibody (mAb) to m-aminophenylboronate (m-APBA) by Flow Microcalorimetry (FMC) and to understand the role of non-specific interactions in the adsorption process. FMC was exploited as a dynamic on-line method to measure instantaneous heat energy transfers in order to understand the surface phenomena underlying mAb’s adsorption towards the synthetic ligand m-APBA under different pH (7.5, 8.5, 9.0, 9.5 and 10.0) and salt concentrations (0 and 150 mM NaCl). Results showed that the adsorption of anti-IL8 mAb to m-APBA is enthalpically driven (ΔH_ads<0), as expected for the predominant reversible esterification reaction between boronates and cis-diols-containing molecules. For all the pH conditions studied, thermograms presented a first exothermic peak, characteristic of the reversible esterification reaction between mAb (pI≥9.3) and m-APBA (pKa = 8.8), except at pH 9.0 in the presence of 150 mM NaCl, for which the thermogram presented a first endothermic peak. The heat of adsorption (ΔH_ads) obtained at conditions where cis-diol interactions were predominant was approximately −243 ± 38 kJ/mol against −82 ± 14 kJ/mol (p-value < 0.05) obtained at pH 9.0 with 150 mM NaCl. The observed shift in the thermogram profile at pH 9.0, 150 mM NaCl, and the consequent decrease of 60–70% in ΔH_ads were indicative of the promotion of electrostatic interactions between the protein and the ligand. Overall, and whereas the binding of the PBA ligand to mAb molecules has been described for decades as being affinity-based, our study demonstrates the multimodal behaviour of this interaction and contributes towards the understanding of the adsorption thermodynamics.

这项工作的目的是调查抗人IL8（抗IL8）单克隆抗体（mAb）的吸附底层到复杂的现象米-aminophenylboronate（米-APBA）用的流动微量（FMC）和理解非特异性相互作用在吸附过程中的作用。FMC被用作动态在线方法来测量瞬时热能传递，以便了解在不同pH（7.5、8.5、9.0、9.5和10.0）和盐浓度下mAb吸附于合成配体m -APBA的潜在表面现象。 （0和150 mM NaCl）。结果表明，抗IL8 mAb与吸附米-APBA是焓驱动（ΔH_广告<0），这是硼酸盐和含顺式二醇的分子之间主要的可逆酯化反应所预期的。对于所研究的所有pH条件，热谱图均显示出第一个放热峰，该峰是mAb（pI≥9.3）和m -APBA（pKa = 8.8）之间可逆酯化反应的特征，但在150 mM NaCl存在下在pH 9.0时除外。热谱图显示了第一个吸热峰。吸附热（ΔH_广告在条件下获得），其中顺式-二醇的相互作用占绝对优势为约-243±针对-82±14千焦/摩尔38千焦/摩尔（p-pH值<0.05），使用150 mM NaCl在pH 9.0下获得。在热分析曲线所观察到的位移，在pH 9.0，150mM的NaCl和的60-70％在ΔH随之减少_广告都说明促进蛋白质和配体之间的静电相互作用的。总体而言，尽管数十年来人们已将PBA配体与mAb分子的结合描述为基于亲和力的，但我们的研究证明了这种相互作用的多峰行为，并有助于理解吸附热力​​学。