RESUMO
INTRODUCTION: Blood proteins are emerging as candidate biomarkers for Alzheimer's disease (AD). We systematically profiled the plasma proteome to identify novel AD blood biomarkers and develop a high-performance, blood-based test for AD. METHODS: We quantified 1160 plasma proteins in a Hong Kong Chinese cohort by high-throughput proximity extension assay and validated the results in an independent cohort. In subgroup analyses, plasma biomarkers for amyloid, tau, phosphorylated tau, and neurodegeneration were used as endophenotypes of AD. RESULTS: We identified 429 proteins that were dysregulated in AD plasma. We selected 19 "hub proteins" representative of the AD plasma protein profile, which formed the basis of a scoring system that accurately classified clinical AD (area under the curve = 0.9690-0.9816) and associated endophenotypes. Moreover, specific hub proteins exhibit disease stage-dependent dysregulation, which can delineate AD stages. DISCUSSION: This study comprehensively profiled the AD plasma proteome and serves as a foundation for a high-performance, blood-based test for clinical AD screening and staging.
Assuntos
Doença de Alzheimer , Peptídeos beta-Amiloides/sangue , Biomarcadores/sangue , Programas de Rastreamento , Proteômica , Proteínas tau/sangue , Idoso , Doença de Alzheimer/sangue , Doença de Alzheimer/diagnóstico , Estudos de Coortes , Endofenótipos , Hong Kong , Humanos , Pessoa de Meia-Idade , Fosforilação , Reprodutibilidade dos TestesRESUMO
In this study, we report a one step quantitative RT-PCR assay for severe acute respiratory syndrome (SARS) diagnosis. The overall detection rate for clinical samples collected from Days 1 to 9 of disease onset is 86.2% and the specificity of the assay is 100%. To detect false negative results due to PCR inhibitors or faulty RNA extraction, we have further modified this one step RT-PCR assay for simultaneous detection of severe acute respiratory syndrome (SARS) coronavirus (CoV) and 18S ribosomal RNA (rRNA) as an internal positive control.
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Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Síndrome Respiratória Aguda Grave/diagnóstico , Coronavírus Relacionado à Síndrome Respiratória Aguda Grave/isolamento & purificação , DNA Ribossômico/análise , Humanos , Nasofaringe/virologia , RNA Ribossômico 18S/genética , RNA Viral/análise , RNA Viral/isolamento & purificação , Padrões de Referência , Coronavírus Relacionado à Síndrome Respiratória Aguda Grave/genética , Sensibilidade e Especificidade , Síndrome Respiratória Aguda Grave/virologiaRESUMO
BACKGROUND: Malaria is one of the most important parasitic infections in humans. A sensitive diagnostic test for malaria that could be applied at the community level could be useful in programs to control the disease. The aim of the present work was to develop a simple, inexpensive molecular test for Plasmodium falciparum. METHODS: Blood was collected from controls (n = 100) and from patients diagnosed with falciparum malaria infection (n = 102), who were recruited to the study. Heat-treated blood samples were tested by a loop-mediated isothermal amplification (LAMP) assay for P. falciparum. Results were interpreted by a turbidity meter in real time or visually at the end of the assay. To evaluate the assay, DNA from these samples was purified and tested by PCR. Results from the LAMP and PCR assays were compared. RESULTS: The LAMP assay detected P. falciparum directly from heat-treated blood. The quantitative data from the assay correlated to the parasite counts obtained by blood-film microscopic analyses. When we used the PCR assay as the comparison method, the sensitivity and specificity of the LAMP assay were 95% and 99%, respectively. CONCLUSIONS: Unlike PCR, the LAMP assay does not require purified DNA for efficient DNA amplification, thereby reducing the cost and turnaround time for P. falciparum diagnosis. The assay requires only basic instruments, and assay positivity can be verified by visual inspection.
Assuntos
DNA de Protozoário/sangue , Temperatura Alta , Malária Falciparum/diagnóstico , Plasmodium falciparum/isolamento & purificação , Reação em Cadeia da Polimerase , Manejo de Espécimes/métodos , Animais , Sequência de Bases , Calibragem , DNA de Protozoário/genética , Humanos , Malária Falciparum/sangue , Dados de Sequência Molecular , Plasmodium falciparum/genética , Reação em Cadeia da Polimerase/economia , Reação em Cadeia da Polimerase/métodos , Reação em Cadeia da Polimerase/normas , Sensibilidade e EspecificidadeRESUMO
We compared the performance of a recently established real-time loop-mediated amplification (LAMP) assay with the one from a highly sensitive quantitative PCR assay. None of these assays produced false-positive results in this study. For samples isolated from patients within the first 3 days of disease onset, the detection rate of the quantitative PCR assay was higher (14 of 15 were positive) than the LAMP assay (9 of 15 were positive). By contrast, the detection rates of these assays toward specimens sampled from patients with more than 3 days of illness were similar (32 of 44 for PCR and 33 of 44 for LAMP were positive). The simpler operation of LAMP might be a possible solution for on-site diagnosis.