Development and Performance Evaluation of A Reagent for Detecting of 17α-hydroxyprogesterone in Dried Blood Spots on Filter Paper Fixed on Blood Spots Stent by Auto TRFIA-4 Automatic Fluorescence Immunoanalyzer
10.3969/j.issn.1671-7414.2025.05.035
- VernacularTitle:Auto TRFIA-4型全自动荧光免疫分析仪血片支架固定滤纸血斑检测17α-羟孕酮的试剂研制及性能评价
- Author:
Yuhua TAN
1
;
Ting YU
;
Haijia YU
;
Meixin CHEN
;
Jingling XIE
;
Jianming FENG
Author Information
1. 广州市丰华生物股份有限公司体外诊断试剂研发中心,广州 510730
- Publication Type:Journal Article
- Keywords:
time-resolving fluorescence immunoassay;
blood spots stent;
filter paper dried blood spots;
17α-hydroxyprogesterone
- From:
Journal of Modern Laboratory Medicine
2025;40(5):182-188
- CountryChina
- Language:Chinese
-
Abstract:
Objective To develop a reagent for detecting of 17α-hydroxyprogesterone(17α-OHP)in dried blood spots on filter paper fixed on vascular stent by Auto TRFIA-4 automatic fluorescence immunoanalyzer and evaluate its performance.Methods The microwell plate was coated with the sheep anti-rabbit IgG antibody as microwell reaction plate,the rabbit anti-human 17α-OHP antibody was diluted as intermediate antibody,and the 17α-OHP-bovine serum albumin conjugate was labeled by europium as europium marker.The concentration of 17α-OHP in dried blood spots on filter paper fixed on blood spots stent was quantitatively detected by Auto TRFIA-4 automatic fluorescence immunoanalyzer.The analysis sensitivity,accuracy,linearity,precision,specificity and stability were evaluated,and whether they met the requirements of the formulated industry standards were evaluated.A total of 227 neonatal heel blood filter paper samples from newborns who were born 72 hours after birth and within 7 days and fully breastfeeding were selected for reagent comparison test.The consistency was analyzed by χ2 test,Kappa test,t test,linear correlation analysis,regression analysis,Bland-Altman method analysis and predictive bias analysis of medical decision level,P<0.05 indicated statistically significant difference.Results The optimal coating concentration of sheep anti-rabbit IgG antibody was 3 μg/ml.The optimal dilution ratio of rabbit anti-human 17α-OHP antibody was 1∶1 500.The optimal dilution ratio of 17α-OHP europium marker mother liquor was 1∶2 500.The limit of blank,limit of detection,limit of quantification was 0.75,1.08 and 1.99 nmol/L,respectively.The relative deviations of the standard check test were within±15.00%,and the average recovery rate was 92.36%.The linear correlation coefficient was 0.997 1 in the range of 2.00 to 300.00nmo/L.The intra-assay and inter-assay coefficients of variation were all within 10.00%.The cross-reactivity rates of 100.00 ng/ml progesterone,17α-hydroxypreg nenolone and 11-deoxycortisol were within 0.089%to 0.64%.The performance of stability test met the requirements.The total coincidence rate was 100%compared with the results of contrast reagent.The quantitative results were highly correlated with the contrast reagent(r=0.999 4,tr=452.02,P<0.05).Conclusion The self-developed reagent has the advantages of high sensitivity,good accuracy,wide linear range,good precision,high specificity and good stability,which meets the requirements of the formulated industry standards,and has high correlation and consistency with the result of contrast reagent,which meets the needs of clinical detection.
