以异硫氰酸苄基乙二胺四乙酸为双功能螯合剂，合成了半抗原，并将半抗原与钥孔戚血蓝素或牛血清白蛋白偶联制备抗原。用二喹啉甲酸(BCA)法测抗原浓度，对半抗原、抗原、KLH和BSA分别进行紫外分光光度计扫描, 利用SDS-PAGE电泳进行定性鉴定，用三硝基苯磺酸法间接测偶联率，用石墨炉原子分光吸收法检测抗原中Hg2+的含量。研究结果表明抗原合成成功， Hg-ITCBE-KLH、Hg-ITCBE-BSA、ITCBE-BSA中载体蛋白ε-氨基的替换程度依次为34.75±4.60％、40.61±0.99％、61.27±0.69％, Hg2+的含量依次为分：38.4±0.5μg/ml、125.5±0.9μg/ml、0μg/ml。
In this paper, artificial hapten Hg-ITCBE was synthesized using ITCBE (Isothiocyanobenzylethyl enediamine tetraacetic acid) as the bifunctional chelating agent. Antigens were synthesized through the coupling the hapten with carrier protein keyhole limpet hemocyanin(KLH) and bovine serum albumin (BSA). After the assay for concentration of the antigen with bicinchoninic acid (BCA) method, the hapten antigen and carrier protein were scanned with ultraviolet spectrophotometer, and verified with SDS-PAGE. The degree of conjugate substitution was quantified using the trintrobenzenesulfonic acid(TNBS) method. Concentration of mercury ion in antigen determined with graphite-furnace atomic absorption spectrometry(AAS). The results indicated antigens were synthesized successfully. The extent of substitution of free lysine residues was 34.75 ± 4.60％ for Hg-ITCBE-KLH, 40.61 ±0.99％for the Hg-ITCBE-BSA and 61.27 ±0.69％ for the ITCBE-BSA. The concentration of mercury ion were 38.4±0.5μg/ml, 125.5±0.9μg/ml and 0μg/ml , respectively.
黄佳俊,王菊芳,刘功良,李志勇. 重金属汞螯合物人工抗原的合成与鉴定[J]. 中国生物工程杂志, 2008, 28(2): 71-75.
. Synthesis and identification of artificial mercury chelate antigen. China Biotechnology, 2008, 28(2): 71-75.
https://manu60.magtech.com.cn/biotech/CN/ 或 https://manu60.magtech.com.cn/biotech/CN/Y2008/V28/I2/71
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