Instituto Grifols S.A., Research & Development Area, Parets del Vallès, Barcelona, Spain.
J Alzheimers Dis. 2012;29(1):159-70. doi: 10.3233/JAD-2012-111139.
Clearance of plasma amyloid-β (Aβ) through plasma exchange and replacement with therapeutic albumin to facilitate net Aβ efflux from the brain to plasma is a novel approach for the treatment of Alzheimer's disease. Therefore, thorough characterization of the capacity of therapeutic albumin to bind Aβ is warranted. In this study, Aβ40 and Aβ42 were quantified by commercial ELISA or Araclon ABtest® in samples of Grifols' therapeutic albumin (Albutein®) 5%, 20%, and 25%. The capacity of Albutein® to bind Aβ was assessed by: a) ELISA in serially diluted therapeutic albumin (0-45 mg/ml protein concentration) to which 80 pg/ml of synthetic Aβ peptide (sAβ40 or sAβ42) were added; b) ELISA in samples of the therapeutic albumin containing serially diluted sAβ40 or sAβ42 (60-400 pg/ml); and c) surface plasmon resonance (SPR) for sAβ42 binding. The Aβ content in Albutein® was below the quantification threshold of the ELISA tests (<25 to <62.5 pg/ml) and ABtest® (<3.125 pg/ml). Quantification of exogenously added sAβ42 decreased in parallel with increasing protein concentration (59-78% at 45 mg/ml albumin). Recovery of sAβ serially diluted in Albutein® was ∼60% for sAβ40 and ∼70% for sAβ42, but was ∼100% in control samples without albumin. The KD by SPR analysis for sAβ42 interaction with Albutein® was 1.72 ± 0.24 × 10-6 M. In conclusion, Grifols' therapeutic albumin has undetectable content of Aβ40 and Aβ42. Moreover, Grifols' therapeutic albumin consistently binds peptides containing the primary sequence of human Aβ.
通过血浆置换清除血浆中的淀粉样蛋白-β (Aβ)，并用治疗性白蛋白替代，以促进 Aβ 从大脑向血浆的净流出，这是治疗阿尔茨海默病的一种新方法。因此，有必要彻底表征治疗性白蛋白结合 Aβ 的能力。在这项研究中，通过商业 ELISA 或 Araclon ABtest®在 Grifols 治疗性白蛋白 (Albutein®) 5%、20%和 25%的样本中定量检测 Aβ40 和 Aβ42。通过以下方法评估 Albutein®结合 Aβ 的能力：a) 在浓度为 0-45mg/ml 蛋白质的连续稀释的 Albutein®中进行 ELISA 实验，其中加入 80pg/ml 的合成 Aβ 肽 (sAβ40 或 sAβ42)；b) 在含有连续稀释 sAβ40 或 sAβ42 的 Albutein®样品中进行 ELISA 实验 (60-400pg/ml)；c) 通过表面等离子体共振 (SPR) 进行 sAβ42 结合实验。Albutein®中的 Aβ 含量低于 ELISA 检测 (<25 至 <62.5pg/ml) 和 ABtest®(<3.125pg/ml)的定量检测限。添加的 sAβ42 的定量随着蛋白质浓度的增加而平行减少 (45mg/ml 白蛋白时为 59-78%)。在 Albutein®中连续稀释的 sAβ 的回收率对于 sAβ40 约为 60%，对于 sAβ42 约为 70%，但在不含白蛋白的对照样品中约为 100%。通过 SPR 分析，sAβ42 与 Albutein®相互作用的 KD 值为 1.72±0.24×10-6M。总之，Grifols 的治疗性白蛋白中检测不到 Aβ40 和 Aβ42 的含量。此外，Grifols 的治疗性白蛋白始终结合含有人类 Aβ 一级序列的肽。
