Lukuan Liu

Multiepitope Templates Imprinted Particles for the Simultaneous Capture of Various Target Proteins

To achieve the simultaneous capture of various target proteins, the multiepitope templates imprinted particles were developed by phase inversion-based poly(ether sulfone) (PES) self-assembly. Herein, with the top three high-abundance proteins in the human plasma, serum albumin, immunoglobulin G, and transferrin, as the target proteins, their N-terminal peptides were synthesized as the epitope templates. After the preorganization of three epitopes and PES in dimethylacetamide, the multiepitope templates imprinted particles were formed in water through self-assembly, by which the simultaneous recognition of three target proteins in human plasma was achieved with high selectivity. Furthermore, the binding kinetics study proved that the adsorption mechanism in this imprinting system toward three epitope templates was the same as that on the single-epitope imprinting polymer. These results demonstrate that our proposed multiepitope templates imprinting strategy might open a new era of artificial antibodies to achieve the recognition of various targets simultaneously.

Epitope imprinting enhanced IMAC (EI-IMAC) for highly selective purification of His-tagged protein

Recombinant protein technology occupies an important position in fields including biopharmaceutics, proteomics, structural and functional biology. However, the purification of His-tagged protein, the majority portion of recombinant protein, is seriously hindered by impurities. These impurities, including host proteins with inherent cysteine and histidine-rich regions or metal centers, are usually beyond the purification ability of commonly used IMAC materials. To remove this barrier, a novel purification material was developed through enhancing the selectivity of IMAC by means of surface epitope imprinting using His-tag, the common terminal of His-tagged protein, as the template. Characterizations including TEM, thermogravimetric analysis, X-ray photoelectron spectroscopy, measurement of DLS size and zeta potential were carried out to prove the fabrication of the imprinted shell. Results exhibited a high imprinting factor of 7.1. Besides, the adsorption kinetics were not affected by the surface imprinted shell and could reach adsorption equilibrium within 15 min. Compared with the substrate IMAC, the novel epitope imprinting enhanced IMAC (EI-IMAC) showed an obvious improvement (5% increase of purity) in the selectivity of His-tagged recombinant protein from crude cell lysis.

Aptamer-immobilized open tubular capillary column to capture circulating tumor cells for proteome analysis

Circulating tumor cells hold the key to predicting the prognosis and discovering the therapeutic targets. Herein, we proposed a strategy to develop an aptamer-immobilized open tubular capillary column by which SMMC-7721 human hepatoma cells (SMMC-7721 cells) could be captured with an over 70% of capture efficiency and a 3.0 ± 0.2 of enrichment factor. Owing to the compatibility of the column, the captured cells by the column could be analyzed by LC-MS from protein level and 5 unique proteins of SMMC-7721 cells were identified which could be used as markers to identify SMMC-7721 cells when Jurkat T-leukemia cells (Jurkat cells) were employed as interfering cells. As the key component, the aptamer-immobilized column had the potential to be integrated into the platform for separating, enriching and characterizing rare cells simultaneously.

Surface sieving coordinated IMAC material for purification of His-tagged proteins

Tailor-made materials for the purification of proteins with His-tag was designed through synergizing the selectivity of surface sieving and metal ion affinity. By excluding impurity proteins out of the surface polymer network, such materials could purify His-tagged proteins from the crude cell lysis with purity up to 90%, improved by 14% compared to that obtained by the commercial metal chelating affinity materials. This study might promote the His-tagged protein purification to a new level.