胶原涂层上化学偶联抗DNA抗体的可行性

Chemical Coupling of Anti-DNA Antibody on Collagen Coating

目的研究抗DNA抗体化学偶联到冠脉支架胶原涂层膜上的可行性及连接的稳定性。方法采用氯胺-T法对抗DNAIgM进行125I标记,将胶原膜随机分为化学偶联组(n=3)和对照组(n=3)J化学偶联组胶原膜用N-琥珀酰亚胺基-3-(2-吡啶基二硫)代丙酸酯(SPDP)通过化学键结合标记IgM,对照组胶原膜通过吸附的方式携带标记的IgM,测定125I的放射活性,评价IgM与胶原膜化学偶联的可行性及稳定性。结果化学偶联组胶原膜上携带的抗体量约为对照组的15倍,与对照组相比差异具有显著性(P<0.01);胶原膜通过化学偶联的方式携带抗体的稳定性明显优于对照组,在37℃60r/min摇动条件下洗脱,对照组胶原膜上携带的抗体仅5d即随胶原膜完全溶解而全部释放,而化学偶联组胶原膜携带的抗体在第16天时仍有25%保留在膜上。结论通过同位素标记定量显示了在蛋白涂层上通过化学偶联的方法连接抗DNA抗体的可行性和稳定性,为下一步冠状动脉血管支架上携带质粒DNA提供了实验依据。

Objective To evaluate the feasibility and stability of chemically conjugating IgM on collagen films. Methods IgM was labeled with ^125I using the chloramine-T method. Six collagen films were randomly divided into two groups. In chemical coupling group ^125I-labelod IgM was chemically coupled with the films through N-succinmiclyl-3- （2-pyridyl-dithio） propionate reaction. In control group ^125I-labelod IgM was absorbed onto collagen films. The amount of IgM on the collagen films and the amount of IgM remained on the films after extensive rinsing with phosphate buffered saline were monitored by counting the radioactivity of ^125I. Results The amount of antibodies loaded onto collagen films in the chemical coupling group was 15 times higher than that on the control films, showing significant statistical difference （P 〈 0.01）. And the stability of conjugation antibodies on collagen films was significantly better than the control films. Conclusion Chemical coupling is an effective approach to immobilize antibodies on collagen for further plasmid DNA tethering.