Department of Biochemistry, Faculty of Biology and Biotechnology, University of Warmia and Mazury, Michała Oczapowskiego 1A, 10-719, Olsztyn, Poland.
Laboratory of Translational Oncology, Intercollegiate Faculty of Biotechnology, Medical University of Gdańsk, Dębinki 1, 80-211, Gdansk, Poland.
J Pharm Biomed Anal. 2021 Mar 20;196:113900. doi: 10.1016/j.jpba.2021.113900. Epub 2021 Jan 17.
Cytokines (CKs) are one of the key components of the molecular network modulating multiple immunological interactions. Within such biological systems, CKs functions are associated with several processes, thus quantification of these analytes in serum samples, as well as a faithful determination of its concentration, are crucial for the translational aspect of many studies.
This study is focused on the evaluation of the effects of storage duration and multiple freeze-thaw cycles on CKs stability.
Serum samples were obtained from 24 healthy participants. Samples were prospectively stored at 4 °C for 1-7 and 30 days, and also underwent multiple freeze-thaw cycles. Afterwards, CK levels were determined by enzyme-linked immunosorbent assay.
Among the 8 examined CKs all of them showed significant degradation (determined with the two-way ANOVA and post-hoc test) after 4 days of sample storage at 4 °C. Serum were affected by freezing at -20 °C and thawing, and 2 of CKs (IL-1β and IL-8) showed significant concentration decrease after following 2 freeze-thaw cycles. It has been also determineded that CKs in serum samples after multiple freeze-thaw cycles had better stability, when samples were stored at -80 °C (compared to storage at -20 °C).
This study demonstrates that long storage at 4 °C, as well as multiple freeze-thaw cycles of serum samples, must be avoided and CK concentrations should be measured immediately after sample collection.
细胞因子（CKs）是调节多种免疫相互作用的分子网络的关键组成部分之一。在这样的生物系统中，CKs 的功能与多个过程相关联，因此，血清样本中这些分析物的定量以及其浓度的准确测定对于许多研究的转化方面至关重要。
本研究旨在评估储存时间和多次冻融循环对 CKs 稳定性的影响。
从 24 名健康参与者中获得血清样本。样本前瞻性地在 4°C 下储存 1-7 天和 30 天，并经历多次冻融循环。随后，通过酶联免疫吸附试验测定 CK 水平。
在所检查的 8 种 CK 中，所有 CK 在 4°C 储存 4 天后均显示出明显的降解（通过双向方差分析和事后检验确定）。血清在-20°C 冷冻和解冻后受到影响，2 种 CK（IL-1β 和 IL-8）在经过 2 次冻融循环后显示出明显的浓度降低。还确定了在多次冻融循环后血清样本中的 CK 具有更好的稳定性，当样本储存在-80°C 时（与储存在-20°C 相比）。
本研究表明，必须避免长时间在 4°C 下储存以及血清样本的多次冻融循环，并且应在样本采集后立即测量 CK 浓度。
