一种综合的自上而下的和自下而上的LC-HRMS/MS方法,用于血和血清中人类生长激素的量化
Sophia Krombholz1, Andreas Thomas1, Philippe Delahaut2
1Center for Preventive Doping Research - Institute of Biochemistry, German Sport University Cologne, Germany.
概括
一种新的液态染色体质谱法 (LC-MS) 方法准确地量化了血和血清中的人类生长激素 (GH). 这种技术为目前的测试提供了有价值的替代方案,并为兴奋剂控制应用奠定了基础.
科学领域:
- 生物化学 生物化学
- 分析化学 分析化学
- 生物技术是生物技术.
背景情况:
- 精确量化人类生长激素 (GH) 对于诊断内分泌疾病,如GH缺乏和壮症至关重要.
- 目前的带结合测定 (LBA) 显示出显著的方法变化,影响诊断可靠性.
- 在临床诊断和兴奋剂控制中需要精确可靠的GH测量方法.
研究的目的:
- 开发和验证一种联合的自上而下和自下而上液态染色体质谱法 (LC-MS) 方法,用于量化血和血清中完整的人类生长激素 (GH).
- 建立未来基于MS的GH在兴奋剂控制中的分析的基础.
- 为了比较开发的LC-MS方法与现有的免疫试验用于GH量化.
主要方法:
- 从血/血清中提取GH通过蛋白沉和使用抗GH抗体和磁珠的免疫捕获.
- 完整的GH使用一个自上而下的2D-LC-HRMS/MS系统进行了分析.
- 样本提取物被消化并使用自下而上的LC-MS分析,以量化特定的GH异型 ('总',22kDa和20kDa).
主要成果:
- 上下LC-HRMS/MS方法准确量化了完整的单体22kDaGH异型在0.5-10 ng/ml的工作范围内.
- 自下而上的分析提供了关于"总"和20kDa的GH异型体的补充数据.
- 来自自上下和自下而上的测试的结果显示,与22kDa异型的常规免疫测试结果有很强的相关性 (R2>0.95).
结论:
- 开发的自上而下的和自下而上的LC-HRMS/MS方法作为一种有价值的补充工具,用于血/血清中的GH量化,增强临床测量.
- 虽然还不足以评估GH滥用,但该方法为未来的LC-MS基GH分析在兴奋剂控制中提供了重要的基础.
- 这种先进的LC-MS方法解决了当前LBA的局限性,提供了更高的精度和更广泛应用的潜力.
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