Section on Clinical Psychoneuroendocrinology and Neuropsychopharmacology, National Institute on Alcohol Abuse and Alcoholism Division of Intramural Clinical and Basic Research and National Institute on Drug Abuse Intramural Research Program, National Institutes of Health, Bethesda, MD, USA.
Eur Rev Med Pharmacol Sci. 2020 Feb;24(3):1585-1589. doi: 10.26355/eurrev_202002_20216.
We evaluated the effect of different concentrations of the esterase inhibitor, AEBSF, and acid treatment on acyl-ghrelin stability in human plasma samples subjected to a freeze/thaw cycle.
Four plasma samples were collected from each donor and treated with the following concentrations of AEBSF: 2 mg/ml, 1 mg/ml, 0.6 mg/ml, and 0 mg/ml. For each plasma tube collected, half of the aliquots were treated with HCl and stored at -80°C before measuring acyl-ghrelin concentration using enzyme-linked immunosorbent assay (ELISA).
Treatment with 1 mg/ml AEBSF + HCl resulted in significantly higher acyl-ghrelin levels compared to all other treatments except 2 mg/ml AEBSF + HCl or 0.6 mg/ml AEBSF + HCl. While all HCl-treated samples had higher acyl-ghrelin levels than their AEBSF-matched un-acidified samples, only samples treated with 1 mg/ml AEBSF significantly differed in acyl-ghrelin levels as a result of HCl treatment.
Our results suggest the use of 1 mg/ml AEBSF with HCl for optimal acyl-ghrelin stability in human plasma samples subjected to a freeze/thaw cycle before assay. Given that 2 mg/ml and 0.6 mg/ml AEBSF + HCl did not significantly differ from 1 mg/ml AEBSF + HCl, our data suggest that the use of AEBSF with HCl more potently prevents de-acylation of ghrelin than either treatment alone.
我们评估了不同浓度的酯酶抑制剂 AEBSF 和酸处理对经受冻融循环的人血浆样品中酰基-ghrelin 稳定性的影响。
从每个供体收集 4 个血浆样品，并分别用以下浓度的 AEBSF 处理：2mg/ml、1mg/ml、0.6mg/ml 和 0mg/ml。对于收集的每个血浆管，一半的等分试样用 HCl 处理，并在 -80°C 下储存，然后使用酶联免疫吸附测定（ELISA）测量酰基-ghrelin 浓度。
用 1mg/ml AEBSF + HCl 处理导致酰基-ghrelin 水平明显高于除 2mg/ml AEBSF + HCl 或 0.6mg/ml AEBSF + HCl 之外的所有其他处理。虽然所有经 HCl 处理的样品的酰基-ghrelin 水平均高于其未经酸化的 AEBSF 匹配样品，但仅用 1mg/ml AEBSF 处理的样品的酰基-ghrelin 水平因 HCl 处理而显著不同。
我们的结果表明，在进行测定之前，对于经受冻融循环的人血浆样品，使用 1mg/ml AEBSF 加 HCl 可最佳稳定酰基-ghrelin。由于 2mg/ml 和 0.6mg/ml AEBSF + HCl 与 1mg/ml AEBSF + HCl 无显著差异，我们的数据表明，AEBSF 加 HCl 的使用比单独处理更有效地防止 ghrelin 的去酰化。
