目的 比较不同分子大小的6A型肺炎球菌(serotype 6A Streptococcus Pneumoniae )结合物和佐剂吸附在小鼠体内免疫原性的影响。方法 通过乙酸水解降低6A型荚膜多糖的相对分子质量制备成水解物,水解物经1-氰基-4-二甲胺基吡啶四氟硼酸盐(C...目的 比较不同分子大小的6A型肺炎球菌(serotype 6A Streptococcus Pneumoniae )结合物和佐剂吸附在小鼠体内免疫原性的影响。方法 通过乙酸水解降低6A型荚膜多糖的相对分子质量制备成水解物,水解物经1-氰基-4-二甲胺基吡啶四氟硼酸盐(CDAP)活化并与破伤风类毒素己二酸酰肼衍生物TT AH 结合,制备成结合物。用Sepharose 4 Fast Flow 纯化结合物,并根据化学检测结果将结合物分为 K D 0.0~0.2、 K D 0.2~0.4、 K D 0.4~0.6等3个组分,每个组分分别以磷酸铝佐剂吸附,将吸附前后的各个组分按照每针次每只小鼠0.2 μg分别免疫小鼠,并采用ELISA检测结合物在小鼠体内的抗体水平。结果 3种不同相对分子质量吸附前后的结合物在小鼠体内均能产生较高水平的抗体,各组2、3针之间具有明显的加强效应。在吸附组和未吸附组中,3种不同分子大小的结合物在小鼠体内产生抗体水平无明显差异。各组分佐剂吸附后的结合物血清抗体滴度高于未吸附组,但这种差异无统计学意义( P > 0.05)。结论 结合物的分子大小对小鼠体内抗体水平的产生没有明显影响;磷酸铝佐剂吸附对于不同分子大小的结合物在小鼠体内的免疫原性有一定的增强效应,但这种增强效应差异无统计学意义。展开更多
Context: Currently, fetal point mutations cannot be reliably analyzed from circulatory fetal DNA in maternal plasma, due to the predominance of maternal DNA sequences. However, analysis of circulatory fetal DNA sequen...Context: Currently, fetal point mutations cannot be reliably analyzed from circulatory fetal DNA in maternal plasma, due to the predominance of maternal DNA sequences. However, analysis of circulatory fetal DNA sequences in maternal plasma have been shown to selectively enrich for fetal DNA molecules on the basis of a smaller molecular size than maternal DNA. Abstract Objective: To examine the prenatal analysis of 4 common β - thalassemia point mutations: IVSI- 1, IVSI- 6, IVSI- 110, and codon 39. Design, Setting, and Patients: A total of 32 maternal blood samples were collected at 10 to 12 weeks of gestation (mean, 10.7 weeks) between February 15, 2003, and February 25, 2004, in Bari, Italy, from women with risk for β - thalassemia in their newborns immediately prior to chorionic villous sampling. Samples in which the father and mother did not carry the same mutation were examined. Circulatory DNA was size- fractionated by gel electrophoresis and polymerase chain reaction (PCR) amplified with a peptide- nucleic- acid clamp, which suppresses amplification of the normal maternal allele. Presence of the paternal mutant allele was detected by allele- specific real- time PCR. Main Outcome Measure: Detection of paternally inherited β - globin gene point mutations. Results: Presence or absence of the paternal mutant allele was correctly determined in 6 (86% ) of 7 cases with the IVSI- 1 mutation, 4 (100% ) of 4 with the IVSI- 6 mutation, 5 (100% ) of 5 with the IVSI- 110 mutation, and 13 (81% ) of 16 with the codon 39 mutation. One false- positive test result was scored for the IVSI- 1 mutation. Two cases with the codon 39 mutation were classified as uncertain and 1 case was excluded due to lack of a diagnostic test result at the time of analysis. These results yielded an overall sensitivity of 100% and specificity of 93.8% , with classified cases removed. Conclusion: Our recently described technique of the size- fractionation of circulatory DNA in maternal plasma may be potentially useful for the noninvasive prenatal determination of fetal point mutations.展开更多
文摘目的 比较不同分子大小的6A型肺炎球菌(serotype 6A Streptococcus Pneumoniae )结合物和佐剂吸附在小鼠体内免疫原性的影响。方法 通过乙酸水解降低6A型荚膜多糖的相对分子质量制备成水解物,水解物经1-氰基-4-二甲胺基吡啶四氟硼酸盐(CDAP)活化并与破伤风类毒素己二酸酰肼衍生物TT AH 结合,制备成结合物。用Sepharose 4 Fast Flow 纯化结合物,并根据化学检测结果将结合物分为 K D 0.0~0.2、 K D 0.2~0.4、 K D 0.4~0.6等3个组分,每个组分分别以磷酸铝佐剂吸附,将吸附前后的各个组分按照每针次每只小鼠0.2 μg分别免疫小鼠,并采用ELISA检测结合物在小鼠体内的抗体水平。结果 3种不同相对分子质量吸附前后的结合物在小鼠体内均能产生较高水平的抗体,各组2、3针之间具有明显的加强效应。在吸附组和未吸附组中,3种不同分子大小的结合物在小鼠体内产生抗体水平无明显差异。各组分佐剂吸附后的结合物血清抗体滴度高于未吸附组,但这种差异无统计学意义( P > 0.05)。结论 结合物的分子大小对小鼠体内抗体水平的产生没有明显影响;磷酸铝佐剂吸附对于不同分子大小的结合物在小鼠体内的免疫原性有一定的增强效应,但这种增强效应差异无统计学意义。
文摘Context: Currently, fetal point mutations cannot be reliably analyzed from circulatory fetal DNA in maternal plasma, due to the predominance of maternal DNA sequences. However, analysis of circulatory fetal DNA sequences in maternal plasma have been shown to selectively enrich for fetal DNA molecules on the basis of a smaller molecular size than maternal DNA. Abstract Objective: To examine the prenatal analysis of 4 common β - thalassemia point mutations: IVSI- 1, IVSI- 6, IVSI- 110, and codon 39. Design, Setting, and Patients: A total of 32 maternal blood samples were collected at 10 to 12 weeks of gestation (mean, 10.7 weeks) between February 15, 2003, and February 25, 2004, in Bari, Italy, from women with risk for β - thalassemia in their newborns immediately prior to chorionic villous sampling. Samples in which the father and mother did not carry the same mutation were examined. Circulatory DNA was size- fractionated by gel electrophoresis and polymerase chain reaction (PCR) amplified with a peptide- nucleic- acid clamp, which suppresses amplification of the normal maternal allele. Presence of the paternal mutant allele was detected by allele- specific real- time PCR. Main Outcome Measure: Detection of paternally inherited β - globin gene point mutations. Results: Presence or absence of the paternal mutant allele was correctly determined in 6 (86% ) of 7 cases with the IVSI- 1 mutation, 4 (100% ) of 4 with the IVSI- 6 mutation, 5 (100% ) of 5 with the IVSI- 110 mutation, and 13 (81% ) of 16 with the codon 39 mutation. One false- positive test result was scored for the IVSI- 1 mutation. Two cases with the codon 39 mutation were classified as uncertain and 1 case was excluded due to lack of a diagnostic test result at the time of analysis. These results yielded an overall sensitivity of 100% and specificity of 93.8% , with classified cases removed. Conclusion: Our recently described technique of the size- fractionation of circulatory DNA in maternal plasma may be potentially useful for the noninvasive prenatal determination of fetal point mutations.
