TrkA-脂筏 @ 琼脂糖凝胶色谱模型的构建及亲和行为研究

打开文本图片集

Construction of TrkA-lipid rafts @ CNBr-Sepharose 4B gels chromatography and its affinity behavior

SHEN Xin，CHI Hao，TANGWeijia，TONG Shanshan （School ofPharmacy，Jiangsu University，ZhenjiangJiangsu 212013）

[Abstract]Objective： To construct a bioaffinity chromatography of TrkA-lipid rafts @ CNBr-Sepharose 4B gels and apply it to the assessment of the efficacy of the affinity adsorption of TrkA targeted drugs.

Methods： TrkA-lipid rafts @ CNBr-Sepharose 4B gels stationary phase was prepared and characterized through the coupling reaction of cyanogen bromide immobilized ligand. The stability was assessed by repeating the up-sampling experiment over a period of 3O days. Affinity adsorption experiments were conducted to investigate the affinity adsorption behavior of TrkA-targeted drugs on both blank agarose gels columns and TrkA-lipid rafts @ CNBr-Sepharose 4B gels columns. Results： The Western blotting results revealed that the lipid rafts contained Flotillin-1，Caveolin-1，and TrkA proteins.Following the coupling of U251 cell lipid rafts coupled Sepharose 4B gels，it was observed that the lipid rafts maintained a stable solid loading on the column bed even after rinsing with multiple column volumes of buffer. Adsorption experiments confirmed the specific affinity adsorption of the TrkA-lipid rafts @ CNBr-Sepharose 4B gels stationary phase. Conclusion： The bioaffinity chromatography of TrkA-lipid rafts @ CNBr-Sepharose 4B gels was successfully constructed， thus preliminarily validating its feasibility and stability.

[Key words] TrkA；lipid rafts；CNBr-Sepharose 4B gels；affnity adsorption；gefitinib

生物亲和色谱作为近年来活性药物筛选的重要技术，其核心优势在于利用固定相上的生物配体与目标分子间的特异性、可逆性相互作用，从而实现对复杂样品中活性成分的高效筛选[1]。（剩余7319字）