化学
免疫球蛋白轻链
单域抗体
抗体
配体(生物化学)
亲和层析
色谱法
洗脱
试剂
噬菌体展示
分子生物学
生物化学
受体
酶
肽
有机化学
免疫学
生物
作者
Zhui Tu,Youlong Xu,Jing-Li Fu,Zhibing Huang,Yao Wang,Bin Liu,Ye Tao
标识
DOI:10.1016/j.jchromb.2014.12.031
摘要
This work aimed to evaluate novel affinity resin used to purify immunoglobulin G (IgG) with a variable domain of the heavy chain of the heavy-chain antibody (VHH) as an affinity ligand. The VHH, isolated from a naïve camelid single-domain phage display library, exhibits not only affinity to the fragment crystallizable (Fc) region of IgG but also high thermal stability. This anti-Fc VHH (AFV) was expressed as a soluble protein in Escherichia coli and purified using a simple heat treatment procedure. The effects of pH and NaCl concentrations on the capacity of AFV resin were also investigated. Results showed a robust property of the AFV resin. It could bind IgGs at various pH conditions (from 6.0 to 9.0) and NaCl concentrations. The static binding capacities of AFV resin ranged from 3.40 ± 0.53 mg/ml to 15.04 ± 0.37 mg/ml measured using rabbit, mouse, and human IgGs. The bound IgGs can be efficiently eluted at pH 5.0, which is conducive to acid-sensitive IgGs and prevents the aggregation of IgGs. After 10 purification cycles or a 7-day period of storage at 37 °C, recovery did not decrease. These findings suggested that VHHs from non-immunized library could also be robust and functional reagent as an affinity purification ligand.
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