Fernández-Sánchez César, McNeil Calum J, Rawson Keith, Nilsson Olle, Leung Hing Y, Gnanapragasam Vincent
School of Clinical and Laboratory Sciences, The Medical School, University of Newcastle upon Tyne, UK.
J Immunol Methods. 2005 Dec 20;307(1-2):1-12. doi: 10.1016/j.jim.2005.08.014. Epub 2005 Oct 24.
The development of a one-step lateral flow immunoassay on a strip format for the rapid and reliable simultaneous detection of serum levels of free and total prostate specific antigen (f-PSA and t-PSA) and estimation of f-PSA to t-PSA ratio (f/t-PSA) is reported. The f/t-PSA ratio has shown to be more specific for the correct diagnosis of prostate cancer than t-PSA alone, especially in the so-called diagnostic grey zone of 4-10 microg/l t-PSA. The performance of the system described relied on non-competitive immunoassay protocols. Herein, f-PSA and t-PSA were sandwiched between anti-f-PSA and anti-t-PSA monoclonal antibodies immobilised on the strip and a colloidal gold anti-PSA antibody tracer. In the presence of PSA in the sample, the tracer accumulated on the strip results in the appearance of specific pink colour lines. The colour intensity of these lines was found to be directly proportional to the PSA concentration and a semi-quantitative estimation could be carried out visually. Quantitative analysis was also possible by densitometry. Using PSA standards prepared in female serum, the strip could be calibrated up to a concentration of 60 microg/l for both PSA species, with an assay time of less than 20 min. The estimated detection limit was 1 microg/l in all cases. The immunostrip showed good storage stability for at least 2 months and the reproducibility was always between 12% and 17%. Fifty-one male serum samples were analysed with the strip and results compared with values obtained by two different commercial immunoassays taken as reference methods. The study yielded acceptable correlation and agreement. An estimation of the sensitivity and specificity demonstrated the strip validity as a potential front-line device for the early detection of prostate cancer and differentiation of benign prostatic anomalies. Small plastic cartridges incorporating the immunostrip and a small blister containing washing solution that helps remove unbound species from the strip were envisaged in order to avoid false positive readings and decrease background signals, thereby leading to better sensitivity and detection limits.
报道了一种基于试纸条形式的一步法侧向流动免疫分析方法，用于快速、可靠地同时检测血清中游离前列腺特异性抗原（f-PSA）和总前列腺特异性抗原（t-PSA）的水平，并估算f-PSA与t-PSA的比值（f/t-PSA）。研究表明，f/t-PSA比值对前列腺癌的正确诊断比单独的t-PSA更具特异性，尤其是在t-PSA为4-10μg/L的所谓诊断灰色区域。所描述系统的性能依赖于非竞争性免疫分析方案。在此，f-PSA和t-PSA夹在固定在试纸上的抗f-PSA和抗t-PSA单克隆抗体与胶体金抗PSA抗体示踪剂之间。当样品中存在PSA时，示踪剂在试纸上的积累会导致出现特定的粉红色条带。发现这些条带的颜色强度与PSA浓度成正比，并且可以通过目视进行半定量估算。也可以通过光密度测定法进行定量分析。使用在女性血清中制备的PSA标准品，该试纸条对两种PSA的校准浓度可达60μg/L，检测时间少于20分钟。在所有情况下，估计的检测限为1μg/L。免疫试纸条显示出至少2个月的良好储存稳定性，重现性始终在12%至17%之间。用该试纸条分析了51份男性血清样本，并将结果与作为参考方法的两种不同商业免疫分析方法获得的值进行了比较。该研究产生了可接受的相关性和一致性。对敏感性和特异性的评估证明了该试纸条作为早期检测前列腺癌和鉴别良性前列腺异常的潜在一线设备的有效性。为了避免假阳性读数并降低背景信号，设想了一种包含免疫试纸条的小塑料盒和一个装有清洗液的小泡罩，清洗液有助于从试纸条上去除未结合的物质，从而提高敏感性和检测限。
