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目的分析新发现的牛蛙凝集素Catesbeianalectin的免疫原性,并筛选其体外特异性结合的糖基配体。方法多肽合成牛蛙凝集素Catesbeianalectin,利用圆二色谱(circular dichroism,CD)检测其二级结构。分别用牛蛙凝集素、麦胚凝集素(wheat germ lectin,WGA)、卵清蛋白(ovalbumin,OVA)免疫BALB/c小鼠,检测血常规与血清效价,分析牛蛙凝集素的免疫原性。表面等离子共振成像(surface plasmon resonance imaging,SPRi)技术筛选30多种与牛蛙凝集素特异性结合的糖基配体。结果牛蛙凝集素Catesbeianalectin为聚脯氨酸2型(PPⅡ)螺旋结构。OVA包被浓度为0.25μg/ml时,小鼠平均血清效价为1∶200;WGA包被浓度为0.008μg/ml时,小鼠平均血清效价为1∶1 600,牛蛙凝集素Catesbeianalectin包被浓度为80μg/ml时,小鼠血清平均效价为1∶25。当30种单糖以浓度5 mmol/L固定于芯片,牛蛙凝集素Catesbeianalectin以浓度0.4 mg/ml通过固定于芯片的糖表面时,主要与半乳糖基和含有半乳糖基的多糖作用。结论牛蛙凝集素Catesbeianalectin免疫原性较低,为S-型凝集素,体外可与带有半乳糖基的多糖结合,为进一步研究牛蛙凝集素Catesbeianalectin的药物靶向载体作用奠定了基础。
OBJECTIVE: To analyze the immunogenicity of the newly discovered bullfrog lectin Catesbeianalectin and to screen its glycosyl-specific ligands in vitro. Methods Peptides synthesized bullfrog lectin Catesbeianalectin and its secondary structure was detected by circular dichroism (CD). BALB / c mice were immunized with bullfrog lectin, wheat germ lectin (WGA) and ovalbumin (OVA) respectively, and the blood routine and serum titer were determined. The immunogenicity of bullfrog lectin was analyzed. Surface plasmon resonance imaging (SPRi) was used to screen more than 30 glycosyl ligands that specifically bind to bullfrog lectins. Results Bullshit lectin Catesbeianalectin is a polyproline type 2 (PP Ⅱ) helical structure. When the OVA coating concentration was 0.25μg / ml, the average serum titer of the mice was 1: 200; when the WGA coating concentration was 0.008μg / ml, the average serum titer of the mice was 1: 1600, and the bullfrog lectin Catesbeianalectin package When the concentration was 80μg / ml, the average mouse serum titer was 1:25. When 30 monosaccharides were immobilized on the chip at a concentration of 5 mmol / L and bullfrog lectin Catesbeianalectin was passed through the sugar surface immobilized on the chip at a concentration of 0.4 mg / ml, galactosyl and galactosyl-containing polysaccharides were mainly used. Conclusion Catesbeianalectin is a low-immunogenicity S-type lectin that binds to galactosyl-containing polysaccharides in vitro, which lays the foundation for further study on the drug-targeted carrier of bullfrog lectin Catesbeianalectin.