Application of Nanotrap technology for high sensitivity measurement of urinary outer surface protein A carboxyl-terminus domain in early stage Lyme borreliosis.
Application of Nanotrap technology for high sensitivity measurement of urinary outer surface protein A carboxyl-terminus domain in early stage Lyme borreliosis.
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DOI：
10.1186/s12967-015-0701-z
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发表时间：
2015-11-04
影响因子：
7.4
通讯作者：
Luchini A
中科院分区：
文献类型：
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作者：
Magni R;Espina BH;Shah K;Lepene B;Mayuga C;Douglas TA;Espina V;Rucker S;Dunlap R;Petricoin EF;Kilavos MF;Poretz DM;Irwin GR;Shor SM;Liotta LA;Luchini A
早期莱姆疏螺旋体病（LB）的及时抗生素治疗可防止进展为严重的多系统疾病。临床上需要在出现强血清学应答之前的时期内提高早期莱姆病测定的诊断特异性。使用一种新的分析物收获纳米技术，Nanotrap颗粒，我们评估了治疗前后早期LB中的尿疏螺旋体外表面蛋白A（OspA）C-末端肽，以及疑似晚期播散性LB的患者。我们采用Nanotrap颗粒浓缩尿OspA，并使用高度特异性的抗OspA单克隆抗体（mAb）作为C-末端肽的检测器。我们将mAb表位映射到狭窄的特异性OspA C-末端结构域OspA 236 -239，其在感染性疏螺旋体物种中保守，但与人蛋白质没有同源性，并且与相关病毒和非疏螺旋体细菌蛋白质没有交叉反应性。在LB流行区收集了268份正在评估所有类型LB的患者的尿液样本。对结果设盲的尿OspA测定利用Nanotrap颗粒预处理、蛋白质印迹法来评估OspA分子大小，并利用OspA肽竞争进行确认。OspA检测特性：灵敏度1.7 pg/mL（最低检测限），%变异系数（CV）= 8%，动态范围1.7-30 pg/mL。治疗前，24/24例新诊断的游走性红斑（EM）皮疹患者尿OspA阳性，而无症状患者的假阳性率为0/117（卡方p < 10−6）。在抗生素治疗过程中表现出EM皮疹持续存在的10例患者中，10/10例为尿OspA阳性。治疗后症状缓解后，8/8例患者的尿OspA从可检测转为不可检测。尿OspA检测对临床症状的特异性为40/40。尿OspA抗原检测对后期血清学结果的特异性为87.5%（21例尿OspA阳性/24例血清学阳性，卡方检验p = 4.072e−15）。100例持续性LB监测患者中41例尿OspA蛋白阳性。OspA尿脱落与并发活动性症状（例如EM皮疹和关节炎）密切相关，而治疗后这些症状的缓解与尿转化为OspA阴性相关。本文的在线版本（doi：10.1186/s12967-015-0701-z）包含补充材料，可供授权用户使用。
Prompt antibiotic treatment of early stage Lyme borreliosis (LB) prevents progression to severe multisystem disease. There is a clinical need to improve the diagnostic specificity of early stage Lyme assays in the period prior to the mounting of a robust serology response. Using a novel analyte harvesting nanotechnology, Nanotrap particles, we evaluated urinary Borrelia Outer surface protein A (OspA) C-terminus peptide in early stage LB before and after treatment, and in patients suspected of late stage disseminated LB. We employed Nanotrap particles to concentrate urinary OspA and used a highly specific anti-OspA monoclonal antibody (mAb) as a detector of the C-terminus peptides. We mapped the mAb epitope to a narrow specific OspA C-terminal domain OspA236-239 conserved across infectious Borrelia species but with no homology to human proteins and no cross-reactivity with relevant viral and non-Borrelia bacterial proteins. 268 urine samples from patients being evaluated for all categories of LB were collected in a LB endemic area. The urinary OspA assay, blinded to outcome, utilized Nanotrap particle pre-processing, western blotting to evaluate the OspA molecular size, and OspA peptide competition for confirmation. OspA test characteristics: sensitivity 1.7 pg/mL (lowest limit of detection), % coefficient of variation (CV) = 8 %, dynamic range 1.7–30 pg/mL. Pre-treatment, 24/24 newly diagnosed patients with an erythema migrans (EM) rash were positive for urinary OspA while false positives for asymptomatic patients were 0/117 (Chi squared p < 10−6). For 10 patients who exhibited persistence of the EM rash during the course of antibiotic therapy, 10/10 were positive for urinary OspA. Urinary OspA of 8/8 patients switched from detectable to undetectable following symptom resolution post-treatment. Specificity of the urinary OspA test for the clinical symptoms was 40/40. Specificity of the urinary OspA antigen test for later serology outcome was 87.5 % (21 urinary OspA positive/24 serology positive, Chi squared p = 4.072e−15). 41 of 100 patients under surveillance for persistent LB in an endemic area were positive for urinary OspA protein. OspA urinary shedding was strongly linked to concurrent active symptoms (e.g. EM rash and arthritis), while resolution of these symptoms after therapy correlated with urinary conversion to OspA negative. The online version of this article (doi:10.1186/s12967-015-0701-z) contains supplementary material, which is available to authorized users.