[Establishment of reference intervals for plasma p-tau181, Aβ1-42, Aβ1-40, and Aβ1-42/40 ratio and a preliminary study of their correlation with corresponding cerebrospinal fluid reference intervals].

Objective: To establish reference intervals for Alzheimer's disease (AD)-related biomarkers in plasma and cerebrospinal fluid (CSF) of adults in Shanghai using single molecule immune detection. Methods: In this cross-sectional study, which included residual plasma samples from 454 healthy individuals and CSF samples from 106 non-AD patients were collected from the Department of Laboratory Medicine, Huashan Hospital, Fudan University, between March and July 2024. The concentrations of phosphorylated tau protein-181 (p-tau181), amyloid-beta peptide 1-42 (Aβ1-42), and amyloid-beta peptide 1-40 (Aβ1-40) were detected using the AST-Dx90 fully automated fluorescence immunoassay analyzer. After normality testing and outlier removal, reference intervals were determined by the non-parametric method and validated using plasma samples from 20 healthy individuals. Results: Spearman correlation analysis revealed no significant correlations with age for plasma Aβ1-42 (R=0.047, P=0.344), the Aβ1-42/40 ratio (R=-0.050, P=0.326), or for CSF p-tau181 (R=-0.078, P=0.438), Aβ1-42 (R=0.002, P=0.980), Aβ1-40 (R=-0.084, P=0.400), and the Aβ1-42/40 ratio (R=0.173, P=0.083) (|R|<0.2, P>0.05). Although plasma p-tau181 (R=-0.102, P=0.032) and Aβ1-40 (R=0.150, P=0.002) showed statistically significant associations with age (P<0.05), the strength of these correlations was very weak (|R|<0.2).Z-tests indicated that there were no statistically significant differences (Z<Z*) in plasma p-tau181 (Z=0.81, Z*=4.08), Aβ1-42 (Z=1.90, Z*=3.94), Aβ1-40 (Z=1.03, Z*=3.96), or the Aβ1-42/40 ratio (Z=0.22, Z*=3.84) between sex subgroups. Similarly, no significant differences were observed for plasma p-tau181 (Z=1.21, Z*=4.08) and Aβ1-40 (Z=3.00, Z*=3.96) between subgroups stratified by the median age (58 years). Therefore, there is no need to establish age-and sex-stratified reference intervals. Consequently, the reference intervals established in this study were as follows: plasma p-tau181≤5.90 pg/ml, Aβ1-42: 26.79-160.30 pg/ml, Aβ1-40: 147.77-501.10 pg/ml, Aβ1-42/40≥0.08; CSF p-tau181≤27.95 pg/ml, Aβ1-42≥179.07 pg/ml, Aβ1-40≤14 466.00 pg/ml, Aβ1-42/40≥0.05. The validation compliance rate for the plasma reference intervals was≥90%. Conclusion: This study successfully established reference intervals for p-tau181, Aβ1-42, Aβ1-40, and Aβ1-42/40 ratio in both plasma and CSF of adults in Shanghai based on single molecule immune detection. 目的： 基于单分子免疫技术建立上海地区成人血浆和脑脊液（CSF）中阿尔茨海默病（AD）相关标志物的参考区间。 方法： 采用横断面研究，共纳入2024年3—7月复旦大学附属华山医院检验医学科454例健康体检人群的剩余血浆样本与106例非AD及其他痴呆患者的CSF样本。采用AST-Dx90全自动荧光免疫分析仪检测磷酸化Tau蛋白-181（p-tau181）、β-淀粉样蛋白1-42（Aβ1-42）、β-淀粉样蛋白1-40（Aβ1-40）浓度。数据经正态性检验、剔除离群值后，采用非参数法确定参考区间，并使用20例健康体检人群的血浆样本验证上述自建参考区间。 结果： Spearman相关分析显示，血浆中Aβ1-42（R=0.047，P=0.344）、Aβ1-42/40比值（R=-0.050，P=0.326）和CSF中p-tau181（R=-0.078，P=0.438）、Aβ1-42（R=0.002，P=0.980）、Aβ1-40（R=-0.084，P=0.400）及Aβ1-42/40比值（R=0.173，P=0.083）与年龄均无显著相关性（|R|<0.2，P>0.05）。血浆p-tau181（R=-0.102，P=0.032）和Aβ1-40（R=0.150，P=0.002）虽与年龄的相关性有统计学意义（P<0.05），但相关性强度极弱（|R|<0.2）。Z检验表明，血浆p-tau181（Z=0.81，Z*=4.08）、Aβ1-42（Z=1.90，Z*=3.94）、Aβ1-40（Z=1.03，Z*=3.96）及Aβ1-42/40比值（Z=0.22，Z*=3.84）在不同性别亚组间，以及血浆p-tau181（Z=1.21，Z*=4.08）、Aβ1-40（Z=3.00，Z*=3.96）按年龄中位数（58岁）分组的亚组间差异均无统计学意义（Z<Z*），因此无需建立年龄性别分层参考区间。最终，本研究建立的参考区间为：血浆p-tau181≤5.90 pg/ml，Aβ1-42为26.79~160.30 pg/ml，Aβ1-40为147.77~501.10 pg/ml，Aβ1-42/40≥0.08；CSF p-tau181≤27.95 pg/ml，Aβ1-42≥179.07 pg/ml，Aβ1-40≤14 466.00 pg/ml，Aβ1-42/40≥0.05。血浆参考区间验证符合率均≥90%。 结论： 本研究基于单分子免疫技术成功建立上海地区成人血浆及CSF中p-tau181、Aβ1-42、Aβ1-40及Aβ1-42/40比值的参考区间。.