Fifth-Generation Digital Immunoassay for Prostate-Specific Antigen by Single Molecule Array Technology
Fifth-Generation Digital Immunoassay for Prostate-Specific Antigen by Single Molecule Array Technology
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DOI：
10.1373/clinchem.2011.169540
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发表时间：
2011-12-01
影响因子：
9.3
通讯作者：
Duffy, David C.
中科院分区：
文献类型：
--
作者：
Wilson, David H.;Hanlon, David W.;Duffy, David C.
背景：现有检测方法的定量限制阻碍了前列腺癌患者根治性前列腺切除术（RP）后前列腺特异性抗原（PSA）的测量。由于根治性前列腺切除术去除了负责产生 PSA 的组织，因此目前的检测方法通常无法检测到术后 PSA。然而，有证据表明，在 RP 后更灵敏地确定 PSA 状态可以改善对患者预后和治疗反应的评估，并为可能受益最大的患者提供更好的针对性二次治疗。我们开发了一种研究性数字免疫测定法，其定量限比当前超灵敏的第三代 PSA 测定法低 2 个对数级。方法：我们开发了用于基于珠子的 ELISA 的试剂，用于飞升体积孔的高密度阵列。将具有免疫复合物和相关酶标记的抗 PSA 捕获珠在阵列的孔内进行单一化，并检查酶产物的存在。我们表征了分析性能，将其与市售测试的准确性进行了比较，并分析了 33 名 RP 患者的初步研究的纵向血清样本。 结果：该测定法表现出功能灵敏度（20% 测定间 CV）< 0.05 pg/mL，从 1 到 50 pg/mL 的总不精密度 < 10%，并且与比较方法具有极好的一致性。所有 RP 样品均在测定测量能力范围内。研究发现手术后的 PSA 浓度可预测 5 年内前列腺癌复发风险。结论：相对于当前的超灵敏测定，定量限度显着提高了 2 个对数，并且该测定经过验证的分析性能允许准确评估 RP 后的 PSA 状态。 (C) 2011 美国临床化学协会
BACKGROUND: Measurement of prostate-specific antigen (PSA) in prostate cancer patients following radical prostatectomy (RP) has been hindered by the limit of quantification of available assays. Because radical prostatectomy removes the tissue responsible for PSA production, postsurgical PSA is typically undetectable with current assay methods. Evidence suggests, however, that more sensitive determination of PSA status following RP could improve assessment of patient prognosis and response to treatment and better target secondary therapy for those who may benefit most. We developed an investigational digital immunoassay with a limit of quantification 2 logs lower than current ultrasensitive third-generation PSA assays.METHODS: We developed reagents for a bead-based ELISA for use with high-density arrays of femtoliter-volume wells. Anti-PSA capture beads with immuno-complexes and associated enzyme labels were singulated within the wells of the arrays and interrogated for the presence of enzymatic product. We characterized analytical performance, compared its accuracy with a commercially available test, and analyzed longitudinal serum samples from a pilot study of 33 RP patients.RESULTS: The assay exhibited a functional sensitivity (20% interassay CV) < 0.05 pg/mL, total imprecision < 10% from 1 to 50 pg/mL, and excellent agreement with the comparator method. All RP samples were well within the assay measurement capability. PSA concentrations following surgery were found to be predictive of prostate cancer recurrence risk over 5 years.CONCLUSIONS: The robust 2-log improvement in limit of quantification relative to current ultrasensitive assays and the validated analytical performance of the assay allow for accurate assessment of PSA status after RP. (C) 2011 American Association for Clinical Chemistry