Establishment and performance evaluations of dot immuno-gold filtration assay for the quantitative determination of serum amyloid A

doi:10.3969/j.issn.1673-8640.2013.09.024

Abstract

Abstract:

Objective To establish a dot immuno-gold filtration assay (DIGFA) for the quantitative determination of serum amyloid A (SAA),and evaluate its analysis performance. Methods Serum DIGFA-based SAA detection reagent (SAA-DOT) was established using 2 anti-SAA monoclonal antibodies being coated with nitrocellulose filter and labeled by colloidal gold, respectively. According to EP files, the SAA-DOT analysis performances (such as detection limit,precision,accuracy,linear range,et al) were evaluated. Results The detection limit of SAA-DOT was 5mg/L. The within-run coefficients of variation (CV) were 9.07% and 10.21% respectively, whereas the inter-day CV were 11.33% and 11.53% respectively, when serum SAA concentrations were 10 and 100 mg/L. The detection linear range was from 5 mg/L to 230 mg/L. SAA-DOT had a good correlationship with latex-enhanced rate nephelometry (LERN) reagent (r=0.995,P<0.05). The median of serum SAA was 5.7 mg/L, and the 95% confidence interval was 0-9.6 mg/L in healthy subjects. Conclusions The DIGFA-based SAA-DOT is convenient, and the reaction can be finished within 5 min. Its performance meets the State Food and Drug Administrration (SFDA) requirements for in vitro diagnostic reagents,which is suitable for the clinical applifcation of serum detection.

Key words: Dot immuno-gold filtration assay, Serum amyloid A, Point-of-care test, Latex-enhanced rate nephelometry

CLC Number:

R446.61

CHEN Changqiang,YANG Jing,GU Zhidong,WAN Yinglei,MAO Kezi,FENG Xiaojing,XU Jianxin,FAN Qishi.. Establishment and performance evaluations of dot immuno-gold filtration assay for the quantitative determination of serum amyloid A[J]. , 2013, 28(9): 835-840.

References

[1] Eklund KK,Niemi K,Kovanen P. Immune functions of serum amyloid A[J] . Crit Rev Immunol,2012,32(4):335-348.
[2] 陈长强. 血清淀粉样蛋白A在疾病应用中的研究进展[J] . 检验医学,2012,27(9): 776-779.
[3] Chan T,Gu F. Early diagnosis of sepsis using serum biomarkers[J] . Expert Rev Mol Diagn,2011,11(5):487-496.
[4] Lannergrd A,Larsson A,Friman G,et al. Human serum amyloid A (SAA) and high sensitive C-reactive protein (hsCRP) in preterm newborn infants with nosocomial infections [J] . Acta Paediatr,2008,97(8):1061-1065.
[5] King VL, Thompson J, Tannock LR. Serum amyloid A in atherosclerosis[J] . Curr Opin Lipidol,2011,22(4):302-307.
[6] Zakynthinos E,Pappa N. Inflammatory biomarkers in coronary artery disease [J] . J Cardiol,2009,53(3):317-333.
[7] Liuzzo G,Biasucci LM,Gallimore JR,et al. The prognostic〖JP〗 value of C-reactive protein and serum amyloid a protein in severe unstable angina [J] . N Engl J Med,1994,331(7): 417-424.
[8] 田小侠,刘乃政,朱桂峰,等. 急性冠状动脉综合征时早期心脏标志物的研究进展[J] . 诊断学理论与实践,2005,4(5): 423-425.
[9] Grebe SO,Kuhlmann U,Fogl D,et al. Macrophage activation is associated with poorer long-term outcomes in renal transplant patients[J] . Clin Transplant,2011,25(5):744-754.
[10] Malle E,Sodin-Semrl S,Kovacevic A. Serum amyloid A: an acute-phase protein involved in tumour pathogenesis [J] . Cell Mol Life Sci,2009,66(1):9-26.
[11] 吕绳凯,刘煜凯. 免疫胶体金的制备及应用[J] . 上海医学检验杂志, 1990,5(1): 62-64.
[12] 曹丹如,张锦锋,刘德琮,等. 快速检测尿微量白蛋白的半定量金免疫渗滤法的研究[J] . 检验医学, 2004,19（5）: 429-431.
[13] Poole S,Walker D,Gaines Das RE,et al. The first international standard for serum amyloid A protein (SAA). Evaluation in an international collaborative study [J] . J Immunol Methods,1998,214(1-2): 1-10.
[14] National Committee for Clinical Laboratory. Protocols for determination of limits of detection and limits of quantitation; approved guideline[S] . EP17-A,NCCLS,2004.
[15] 冯仁丰. 临床检验质量管理技术基础[M] . 第2版. 上海: 上海科学技术文献出版社,2007: 50-65.
[16]National Committee for Clinical Laboratory. Evaluation of precision performance of quantitative measurement methods; approved guideline[S] . EP5-A2,NCCLS,2004.[17] National Committee for Clinical Laboratory. Method comparison and bias estimation using patient samples; approved guideline[S] . EP9-A2,NCCLS,2002.
[18] National Committee for Clinical Laboratory. Evaluation of the linearity of quantitative measurement procedures: a statistical approach; approved guideline[S] . EP6-A,NCCLS,2004.
[19] Clinical and Laboratory Standards Institute. Defining,establishing,and verifying reference intervals in the clinical laboratory; approved guideline[S] . C28-A3,CLSI,2008.
[20] Rosenthal CJ,Franklin EC. Variation with age and disease of an amyloid A protein-related serum component[J] . J Clin Invest, 1975,55(4): 746-753.