Liang Kai, Wu Hongmei, Hu Tony Y, Li Yan
Laboratory of Interdisciplinary Research, Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101 China.
Laboratory of Interdisciplinary Research, Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101 China ; GuangDong Bio-healtech Advanced Co., Ltd, Foshan City, 52800 GuangDong Province China.
Clin Proteomics. 2016 Nov 22;13:34. doi: 10.1186/s12014-016-9134-9. eCollection 2016.
High quality clinical samples are critical for meaningful interpretation of data obtained in both basic and translational medicine. More specifically, optimized pre-analysis handling to bio-sample is crucial for avoiding biased analysis in a clinical setting. A universally applicable method for the evaluation of sample quality and pre-analysis handling is therefore in great demand.
The fingerprint pattern of low molecular weight (LMW) peptides in sera is directly associated with sample quality and handling process. Previous studies for enrichment/isolation of LMW peptides have shown that LMW peptides can be enriched by silica meso-porous material in a sensitive and high-throughput manner. Here, a peptide profile approach utilizing mesoporous silica chip-based sample preparation combined with MALDI MS analysis was used as a new platform for evaluation of bio-sample quality. Rat sera were selected as model sample and analyzed according to their LMW peptide fingerprint spectra.
This novel method can complete the entire sample preparation procedure in a short period of time (<40 min), requires minimum amounts of sample (<10 µL), is of high sensitivity (LOD 10 ng/mL) as well as high reproducibility (CV% < 15%). According to the acquired LMW peptide spectra, we were able to distinguish the serum samples processed under different conditions (including different storage temperature, time, and freezing/thaw cycles) with the help of bioinformatics tools (principle composition analysis and significant difference analysis), and identify the samples that had significantly changed due to the inappropriate processing. Based on the percentage of significantly changed peaks in LMW peptide mass spectrum after handling, a judgment standard was established that can be used to evaluate the status of preservation of a biological sample. In addition, our principle study established recommendations for storage time, storage temperature and freeze/thaw conditions.
Our novel method for analysis of bio-samples allows for effective identification of variations in composition within samples, and provides a cost-effective tool for simple sample manipulation in a clinical setting.
高质量的临床样本对于在基础医学和转化医学中获得的数据进行有意义的解读至关重要。更具体地说，对生物样本进行优化的分析前处理对于避免临床环境中的偏倚分析至关重要。因此，迫切需要一种普遍适用的样本质量评估和分析前处理方法。
血清中低分子量（LMW）肽的指纹图谱与样本质量和处理过程直接相关。先前关于LMW肽富集/分离的研究表明，LMW肽可以通过介孔二氧化硅材料以灵敏且高通量的方式进行富集。在此，一种利用基于介孔二氧化硅芯片的样本制备结合基质辅助激光解吸电离质谱（MALDI MS）分析的肽谱方法被用作评估生物样本质量的新平台。选择大鼠血清作为模型样本，并根据其LMW肽指纹图谱进行分析。
这种新方法能够在短时间内（<40分钟）完成整个样本制备过程，所需样本量最少（<10μL），具有高灵敏度（检测限10 ng/mL）以及高重现性（变异系数<15%）。根据获得的LMW肽谱，我们能够借助生物信息学工具（主成分分析和显著性差异分析）区分在不同条件下（包括不同储存温度、时间和冻融循环）处理的血清样本，并识别出因处理不当而发生显著变化的样本。基于处理后LMW肽质谱中显著变化峰的百分比，建立了一个可用于评估生物样本保存状态的判断标准。此外，我们的初步研究确定了储存时间、储存温度和冻融条件的建议。
我们用于分析生物样本的新方法能够有效识别样本内成分的变化，并为临床环境中的简单样本处理提供了一种经济高效的工具。
