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1.
Int J Mol Sci ; 24(24)2023 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-38139429

RESUMO

Diabetes nephropathy (DN) is the leading cause of end-stage renal disease (ESRD) worldwide, and podocyte injury is the central contributor to the progression of DN. Despite the emerging evidence that has established the importance of podocyte endoplasmic reticulum (ER) stress in the pathogenesis of DN, abnormal protein O-GlcNAcylation is also augmented. Currently, the mechanism associating these two hyperglycemia-induced disorders remains poorly understood. This study intended to elucidate whether ER stress drives hyper-protein O-GlcNAcylation to cause podocyte injury in DN. We used both type 1 and type 2 DN models to confirm the occurrence of ER stress and excessive protein O-GlcNAcylation, and then podocyte purification was also conducted for further investigation. Nephroseq V5 data were mined and in vitro studies were applied to reveal the involvement of ER stress and hyper-O-GlcNAcylation in podocyte injury. Our results indicated that ER stress was induced in both type 1 and type 2 DN, and the human RNA-seq data from Nephroseq V5 showed that O-GlcNAcylation-related genes were significantly upregulated in the DN patients. We further demonstrated that ER stress occurred prior to hyper-O-GlcNAc modification and that pharmacologically inhibited protein O-GlcNAcylation can help decrease the podocyte apoptosis induced by hyperglycemia. Together, these discoveries will aid in uncovering the activation of the ER stress-O-GlcNAcylation axis in podocyte injury under DN, which will help open up new therapeutic approaches for preventing DN progression.


Assuntos
Diabetes Mellitus Tipo 2 , Nefropatias Diabéticas , Hiperglicemia , Podócitos , Humanos , Podócitos/metabolismo , Nefropatias Diabéticas/metabolismo , Estresse do Retículo Endoplasmático/genética , Proteínas/metabolismo , Hiperglicemia/metabolismo , Diabetes Mellitus Tipo 2/metabolismo
2.
Cells ; 10(10)2021 09 28.
Artigo em Inglês | MEDLINE | ID: mdl-34685560

RESUMO

The increasing load of senescent cells is a source of aging, and chronic inflammation plays a pivotal role in cellular senescence. In addition, senescent renal tubular epithelial cells are closely associated with renal aging. Lysophosphatidic acid (LPA) is a bioactive lipid mainly produced by the catalytic action of autotaxin (ATX), and its ligation to LPA receptor-1 (LPAR1) is associated with chronic inflammation and renal fibrosis; however, its role in renal aging is unclear. Male 2-, 12-, and 24-month-old C57BL/6 mice and Human renal proximal tubular epithelial cells (HRPTEpiC) were used in the present study. DNA damage and oxidative stress-induced senescence were simulated using doxorubicin (DOXO) and H2O2, respectively. The aged kidney showed decreased renal function, increased fractional mesangial area, and tubulointerstitial fibrosis. Both aged kidney and senescent cells showed increased levels of LPAR1, Nuclear factor κB (NF-κB), and inflammatory cytokines. In addition, LPAR1-knockdown reduced NF-κB and subsequent inflammatory cytokine induction, and NF-κB-knockdown resulted in decreased LPAR1 expression. Our study revealed a positive feedback loop between LPAR1 and NF-κB, which reinforces the role of inflammatory response, suggesting that blocking of aberrantly activated LPAR1 may reduce excessive inflammation, thereby providing a new possible therapeutic strategy to attenuate renal aging.


Assuntos
Inflamação/genética , Nefropatias/genética , Receptores de Ácidos Lisofosfatídicos/metabolismo , Envelhecimento , Animais , Humanos , Masculino , Camundongos , Transdução de Sinais
3.
Nat Commun ; 12(1): 1025, 2021 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-33589623

RESUMO

A black hole X-ray binary produces hard X-ray radiation from its corona and disk when the accreting matter heats up. During an outburst, the disk and corona co-evolves with each other. However, such an evolution is still unclear in both its geometry and dynamics. Here we report the unusual decrease of the reflection fraction in MAXI J1820+070, which is the ratio of the coronal intensity illuminating the disk to the coronal intensity reaching the observer, as the corona is observed to contrast during the decay phase. We postulate a jet-like corona model, in which the corona can be understood as a standing shock where the material flowing through. In this dynamical scenario, the decrease of the reflection fraction is a signature of the corona's bulk velocity. Our findings suggest that as the corona is observed to get closer to the black hole, the coronal material might be outflowing faster.

4.
Adv Mater ; 28(36): 7929-7935, 2016 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-27376951

RESUMO

A novel biomimetic immuno-magnetosome (IMS) is developed by coating a leukocyte membrane (decorated with anti-epithelial cell-adhesion molecule antibody) on a magnetic nanocluster. In addition to the good stability and magnetic controllability, the IMS also exhibits satisfactory binding avidity to circulating tumor cells but stealth property to leukocytes. As a result, rare tumor cells can be effectively enriched with undetectable leukocyte background.


Assuntos
Materiais Biomiméticos , Separação Celular/instrumentação , Separação Celular/métodos , Magnetossomos/imunologia , Células Neoplásicas Circulantes , Moléculas de Adesão Celular/imunologia , Contagem de Células , Humanos , Leucócitos/citologia
5.
Analyst ; 141(10): 2948-54, 2016 05 10.
Artigo em Inglês | MEDLINE | ID: mdl-27049139

RESUMO

Ru(ii) polypyridyl complexes have been expected as promising therapeutic agents against cancer owing to its DNA photocleavage activity. However, the lack of cell selectivity poses a significant obstacle to their practical application. Herein, the strategy combining cell-specific imaging with photoinduced cell death based on [Ru(phen)2(dppz)](2+) has been developed by incorporating [Ru(phen)2(dppz)](2+) into folate-conjugated liposomes. The cells overexpressing folate receptors could specifically recognize this vehicle and be imaged through the luminescence of [Ru(phen)2(dppz)](2+). Thereafter, the delivered [Ru(phen)2(dppz)](2+) interacted with DNA in cells and led to photoinduced cell death. This work provided a possible alternative for cancer diagnosis and therapy.


Assuntos
Ácido Fólico/química , Compostos Organometálicos/química , Rutênio/química , Células A549 , Morte Celular , DNA , Células HeLa , Humanos , Lipossomos , Luminescência , Células MCF-7 , Fototerapia
6.
BMC Cancer ; 14: 747, 2014 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-25282590

RESUMO

BACKGROUND: We investigated the role of the HIPK2-p53 signaling pathway in tumorigenesis and resistance to the drug Verbascoside (VB) in colorectal cancer (CRC), using in vivo and in vitro experiments. METHODS: Primary human CRC samples and normal intestinal tissues from patients were analyzed for HIPK2 expression by immunohistochemistry (IHC) and its expression was correlated against patients' clinicopathological characteristics. Human CRC HCT-116 cells were implanted in BALB/c nude mice; mice with xenografted tumors were randomly administrated vehicle (control), 20, 40, or 80 mg/mL VB, or 1 mg/mL fluorouracil (5-FU). HIPK2, p53, Bax, and Bcl-2 expression in these tumors were determined by IHC. In vitro effects of VB on CRC cell proliferation and apoptosis were measured by CCK-8 assay and flow cytometry; HIPK2, p53, p-p53, Bax, and Bcl-2 were measured by western blot. RESULTS: IHC analysis for 100 human CRC tumor samples and 20 normal intestinal tissues, showed HIPK2 expression to inversely correlate with Dukes stage and depth of invasion in CRC (P<0.05). In vivo, the inhibition rates of 20, 40, and 80 mg/mL VB on CRC xenograft tumor weight were 42.79%, 53.90%, and 60.99%, respectively, and were accompanied by increased expression of HIPK2, p53, and Bax, and decreased Bcl-2 expression in treated tumors. In vitro, VB significantly inhibited proliferation of CRC cell lines HCT-116, HT-29, LoVo, and SW620, in a time- and dose-dependent manner. The apoptosis rates of 25, 50, and 100 µM VB on HCT-116 cells were 10.83±1.28, 11.25±1.54, and 20.19±2.87%, and on HT-29 cells were 18.92±6.12, 21.57±4.05, and 25.14±6.73%, respectively. In summary, VB treatment significantly enhanced the protein expression of pro-apoptotic HIPK2, p53, p-p53, Bax, and decreased anti-apoptotic Bcl-2 expression in CRC cells. CONCLUSIONS: HIPK2 protein modulates the phosphorylation status of p53, and levels of Bax and Bcl-2 in CRC. We also found that VB effectively activated the HIPK2-p53 signaling pathway, resulting in increased CRC cell apoptosis.


Assuntos
Apoptose/efeitos dos fármacos , Proteínas de Transporte/metabolismo , Neoplasias Colorretais/metabolismo , Glucosídeos/farmacologia , Fenóis/farmacologia , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteína Supressora de Tumor p53/metabolismo , Adulto , Idoso , Animais , Antineoplásicos Fitogênicos/farmacologia , Apoptose/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Modelos Animais de Doenças , Feminino , Expressão Gênica , Xenoenxertos , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Gradação de Tumores , Metástase Neoplásica , Estadiamento de Neoplasias
7.
PLoS One ; 9(2): e88484, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24586333

RESUMO

Recently published international guidelines recommend the clinical use of noninvasive prenatal test (NIPT) for aneuploidy screening only among pregnant women whose fetuses are deemed at high risk. The applicability of NIPT to aneuploidy screening among average risk pregnancies requires additional supportive evidence. A key determinant of the reliability of aneuploidy NIPT is the fetal DNA fraction in maternal plasma. In this report, we investigated if differences in fetal DNA fractions existed between different pregnancy risk groups. One hundred and ninety-five singleton pregnancies with male fetuses divided into 3 groups according to first trimester screening parameters were examined for fetal DNA percentage by counting Y chromosome DNA sequences using massively parallel sequencing. Fetal DNA fractions were compared between risk groups and assessed for correlations with first trimester screening parameters. There was no statistically significant difference in fetal DNA fractions across the high, intermediate and low risk groups. Fetal DNA fraction showed a strong negative correlation with maternal weight. Fetal DNA fraction also showed weak but significant correlations with gestational age, crown-rump length, multiple of medians of free ß-subunit of human chorionic gonadotropin and pregnancy-associated plasma protein A. Similar fetal DNA fractions in maternal plasma between high, intermediate and low risk pregnant women is a precondition for uniform performance of the aneuploidy NIPTs for the general population. This study thus shows that the aneuploidy screening by NIPT is likely to offer similar analytical reliability without respect to the a priori fetal aneuploidy risk.


Assuntos
Aneuploidia , DNA/genética , Cromossomos Humanos Y/genética , Feminino , Feto/metabolismo , Humanos , Masculino , Gravidez , Primeiro Trimestre da Gravidez/genética , Diagnóstico Pré-Natal/métodos
8.
Anal Bioanal Chem ; 406(11): 2687-93, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24500758

RESUMO

The sialic acid (SA)-phenylboronic acid (PBA) recognition system is of particular interest in the bioconjugation field, because it is simple, fast, efficient, and biocompatible. In this paper, we report a novel method for reversibly labeling living virus with quantum dots (QDs) by taking advantage of this SA-PBA recognition system. The QDs were initially modified with PBA (QDs-PBA) to target them to the surface of vesicular stomatitis virus (VSV), which has abundant with SA on its envelope. The QDs-PBA was of good monodispersity and strong fluorescence, and could be conjugated with VSV by simply incubating with native VSV for 10 min at 37 °C, producing QDs-VSV that was capable of being imaged at the single virion level. The labeling efficiency attained 83 ± 4.3 % (mean ± SD); meanwhile, the activity and recognition ability of the labeled virus were minimally affected. This method was simple, rapid, and reversible. This work promotes the virus labeling development to a new step. That is, native viruses can be reversibly labeled without any modification.


Assuntos
Ácidos Borônicos/química , Ácido N-Acetilneuramínico/química , Coloração e Rotulagem/métodos , Vírus/química , Pontos Quânticos
9.
PLoS One ; 6(7): e21791, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-21755002

RESUMO

Massively parallel sequencing of DNA molecules in the plasma of pregnant women has been shown to allow accurate and noninvasive prenatal detection of fetal trisomy 21. However, whether the sequencing approach is as accurate for the noninvasive prenatal diagnosis of trisomy 13 and 18 is unclear due to the lack of data from a large sample set. We studied 392 pregnancies, among which 25 involved a trisomy 13 fetus and 37 involved a trisomy 18 fetus, by massively parallel sequencing. By using our previously reported standard z-score approach, we demonstrated that this approach could identify 36.0% and 73.0% of trisomy 13 and 18 at specificities of 92.4% and 97.2%, respectively. We aimed to improve the detection of trisomy 13 and 18 by using a non-repeat-masked reference human genome instead of a repeat-masked one to increase the number of aligned sequence reads for each sample. We then applied a bioinformatics approach to correct GC content bias in the sequencing data. With these measures, we detected all (25 out of 25) trisomy 13 fetuses at a specificity of 98.9% (261 out of 264 non-trisomy 13 cases), and 91.9% (34 out of 37) of the trisomy 18 fetuses at 98.0% specificity (247 out of 252 non-trisomy 18 cases). These data indicate that with appropriate bioinformatics analysis, noninvasive prenatal diagnosis of trisomy 13 and trisomy 18 by maternal plasma DNA sequencing is achievable.


Assuntos
Cromossomos Humanos Par 13/genética , Cromossomos Humanos Par 18/genética , DNA/sangue , Feto/patologia , Diagnóstico Pré-Natal/métodos , Análise de Sequência de DNA , Trissomia/diagnóstico , Composição de Bases/genética , Feminino , Genoma Humano/genética , Humanos , Gravidez , Trissomia/genética
10.
Clin Chem ; 54(3): 500-11, 2008 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-18202156

RESUMO

BACKGROUND: The presence of fetal DNA in maternal plasma represents a source of fetal genetic material for noninvasive prenatal diagnosis; however, the coexisting background maternal DNA complicates the analysis of aneuploidy in such fetal DNA. Recently, the SERPINB5 gene on chromosome 18 was shown to exhibit different DNA-methylation patterns in the placenta and maternal blood cells, and the allelic ratio for placenta-derived hypomethylated SERPINB5 in maternal plasma was further shown to be useful for noninvasive detection of fetal trisomy 18. METHODS: To develop a similar method for the noninvasive detection of trisomy 21, we used methylation-sensitive single nucleotide primer extension and/or bisulfite sequencing to systematically search 114 CpG islands (CGIs)-76% of the 149 CGIs on chromosome 21 identified by bioinformatic criteria-for differentially methylated DNA patterns. The methylation index (MI) of a CpG site was estimated as the proportion of molecules methylated at that site. RESULTS: We identified 22 CGIs which were shown to contain CpG sites that were either completely unmethylated (MI = 0.00) in maternal blood cells and methylated in the placenta (MI range, 0.22-0.65), or completely methylated (MI = 1.00) in maternal blood cells and hypomethylated in the placenta (MI range, 0.00-0.75). We detected, for the first time, placental DNA-methylation patterns on chromosome 21 in maternal plasma during pregnancy and observed their postpartum clearance. CONCLUSION: Twenty-two (19%) of the 114 studied CGIs on chromosome 21 showed epigenetic differences between samples of placenta and maternal blood cells; these CGIs may provide a rich source of markers for noninvasive prenatal diagnosis.


Assuntos
Cromossomos Humanos Par 21/genética , Metilação de DNA , Síndrome de Down/diagnóstico , Epigênese Genética , Placenta/metabolismo , Diagnóstico Pré-Natal/métodos , Biomarcadores/sangue , Ilhas de CpG , Feminino , Feto , Marcadores Genéticos , Humanos , Plasma , Período Pós-Parto , Gravidez
11.
Clin Chem ; 53(12): 2205-9, 2007 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-17951288

RESUMO

BACKGROUND: The specific detection of a minor population of mutant DNA molecules requires methods of high specificity and sensitivity. While the single-allele base extension reaction (SABER) was shown to be useful for the detection of certain beta-thalassemia mutations, we encountered problems with false positivity during development of SABER for the noninvasive prenatal diagnosis of the hemoglobin E (HbE) disease. Systematic optimization resulted in an alternative protocol, the allele-specific base extension reaction (ASBER). METHODS: An artificial model was established by mixing genomic DNA of HbE carriers and normal individuals. Effects of terminator concentration and annealing temperature on the nonspecificity of SABER were then studied. The use of a single relevant terminator and the other 3 types of dideoxynucleotide as competing terminators were also compared in the development of the ASBER protocol. Thirteen cases of HbE-susceptible pregnancies were tested to compare the SABER and the ASBER protocols. RESULTS: Decreasing the single relevant terminator concentration and increasing the annealing temperature in SABER were found to improve specificity. The use of the other 3 types of dideoxynucleotide as competing terminators was shown to offer better detection sensitivity than a single terminator in ASBER. Genotyping results were all correctly determined by ASBER, except one false-negative detection (sensitivity: 80%, specificity: 100%). CONCLUSIONS: An alternative mass spectrometry-based protocol for noninvasive prenatal diagnosis, ASBER, has been successfully developed to allow the detection of a minor DNA population with a point mutation.


Assuntos
Hemoglobina E/genética , Hemoglobinopatias/diagnóstico , Diagnóstico Pré-Natal/métodos , Alelos , Reações Falso-Positivas , Feminino , Genótipo , Testes Hematológicos , Heterozigoto , Humanos , Espectrometria de Massas , Mutação Puntual , Gravidez , Sensibilidade e Especificidade
12.
Nat Med ; 13(2): 218-23, 2007 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-17206148

RESUMO

Current methods for prenatal diagnosis of chromosomal aneuploidies involve the invasive sampling of fetal materials using procedures such as amniocentesis or chorionic villus sampling and constitute a finite risk to the fetus. Here, we outline a strategy for fetal chromosome dosage assessment that can be performed noninvasively through analysis of placental expressed mRNA in maternal plasma. We achieved noninvasive prenatal diagnosis of fetal trisomy 21 by determining the ratio between alleles of a single-nucleotide polymorphism (SNP) in PLAC4 mRNA, which is transcribed from chromosome 21 and expressed by the placenta, in maternal plasma. PLAC4 mRNA in maternal plasma was fetal derived and cleared after delivery. The allelic ratios in maternal plasma correlated with those in the placenta. Fetal trisomy 21 was detected noninvasively in 90% of cases and excluded in 96.5% of controls.


Assuntos
Cromossomos Humanos Par 21/genética , Embrião de Mamíferos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Placenta/metabolismo , Diagnóstico Pré-Natal/métodos , RNA/sangue , Trissomia/genética , Povo Asiático/genética , Feminino , Frequência do Gene , Humanos , Polimorfismo de Nucleotídeo Único , Gravidez , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , População Branca/genética
13.
Biosci Biotechnol Biochem ; 70(9): 2262-7, 2006 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-16960371

RESUMO

Two novel genes (tsB, tsC) involved in the conversion of DL-2-amino-Delta2-thiazoline-4-carboxylic acid (DL-ATC) to L-cysteine through S-carbamyl-L-cysteine (L-SCC) pathway were cloned from the genomic DNA library of Pseudomonas sp. TS1138. The recombinant proteins of these two genes were expressed in Escherichia coli BL21, and their enzymatic activity assays were performed in vitro. It was found that the tsB gene encoded an L-ATC hydrolase, which catalyzed the conversion of L-ATC to L-SCC, while the tsC gene encoded an L-SCC amidohydrolase, which showed the catalytic ability to convert L-SCC to L-cysteine. These results suggest that tsB and tsC play important roles in the L-SCC pathway and L-cysteine biosynthesis in Pseudomonas sp. TS1138, and that they have potential applications in the industrial production of L-cysteine.


Assuntos
Cisteína/análogos & derivados , Cisteína/metabolismo , Genes Bacterianos , Pseudomonas/genética , Pseudomonas/metabolismo , Tiazóis/metabolismo , Sequência de Bases , Cromatografia em Camada Fina , Clonagem Molecular , DNA Bacteriano/química , DNA Bacteriano/genética , Eletroforese em Gel de Poliacrilamida , Biblioteca Gênica , Microbiologia Industrial/métodos , Dados de Sequência Molecular , Pseudomonas/enzimologia , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Análise de Sequência de DNA
14.
BMC Infect Dis ; 6: 20, 2006 Feb 09.
Artigo em Inglês | MEDLINE | ID: mdl-16466582

RESUMO

BACKGROUND: We have previously developed a test for the diagnosis and prognostic assessment of the severe acute respiratory syndrome (SARS) based on the detection of the SARS-coronavirus RNA in serum by real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR). In this study, we evaluated the feasibility of automating the serum RNA extraction procedure in order to increase the throughput of the assay. METHODS: An automated nucleic acid extraction platform using the MagNA Pure LC instrument (Roche Diagnostics) was evaluated. We developed a modified protocol in compliance with the recommended biosafety guidelines from the World Health Organization based on the use of the MagNA Pure total nucleic acid large volume isolation kit for the extraction of SARS-coronavirus RNA. The modified protocol was compared with a column-based extraction kit (QIAamp viral RNA mini kit, Qiagen) for quantitative performance, analytical sensitivity and precision. RESULTS: The newly developed automated protocol was shown to be free from carry-over contamination and have comparable performance with other standard protocols and kits designed for the MagNA Pure LC instrument. However, the automated method was found to be less sensitive, less precise and led to consistently lower serum SARS-coronavirus concentrations when compared with the column-based extraction method. CONCLUSION: As the diagnostic efficiency and prognostic value of the serum SARS-CoV RNA RT-PCR test is critically associated with the analytical sensitivity and quantitative performance contributed both by the RNA extraction and RT-PCR components of the test, we recommend the use of the column-based manual RNA extraction method.


Assuntos
Técnicas de Diagnóstico Molecular/métodos , RNA Viral/isolamento & purificação , Síndrome Respiratória Aguda Grave/diagnóstico , Coronavírus Relacionado à Síndrome Respiratória Aguda Grave/isolamento & purificação , Humanos , RNA Viral/sangue , Kit de Reagentes para Diagnóstico , Análise de Regressão , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Coronavírus Relacionado à Síndrome Respiratória Aguda Grave/genética , Sensibilidade e Especificidade , Síndrome Respiratória Aguda Grave/virologia , Carga Viral
15.
BMC Infect Dis ; 5: 87, 2005 Oct 18.
Artigo em Inglês | MEDLINE | ID: mdl-16229749

RESUMO

BACKGROUND: The Severe Acute Respiratory Syndrome (SARS) was a newly emerged infectious disease which caused a global epidemic in 2002-2003. Sequence analysis of SARS-coronavirus isolates revealed that specific genotypes predominated at different periods of the epidemic. This information can be used as a footprint for tracing the epidemiology of infections and monitor viral evolution. However, direct sequencing analysis of a large number of clinical samples is cumbersome and time consuming. We present here a simple and rapid assay for the screening of SARS-coronavirus genotypes based on the use of fluorogenic oligonucleotide probes for allelic discrimination. METHODS: Thirty SARS patients were recruited. Allelic discrimination assays were developed based on the use of fluorogenic oligonucleotide probes (TaqMan). Genotyping of the SARS-coronavirus isolates obtained from these patients were carried out by the allelic discrimination assays and confirmed by direct sequencing. RESULTS: Genotyping based on the allelic discrimination assays were fully concordant with direct sequencing. All of the 30 SARS-coronavirus genotypes studied were characteristic of genotypes previously documented to be associated with the latter part of the epidemic. Seven of the isolates contained a previously reported major deletion but in patients not epidemiologically related to the previously studied cohort. CONCLUSION: We have developed a simple and accurate method for the characterization and screening of SARS-coronavirus genotypes. It is a promising tool for the study of epidemiological relationships between documented cases during an outbreak.


Assuntos
Evolução Molecular , Síndrome Respiratória Aguda Grave/virologia , Coronavírus Relacionado à Síndrome Respiratória Aguda Grave/genética , Alelos , Genes Virais/genética , Genótipo , Humanos , Sondas de Oligonucleotídeos/análise , Sondas de Oligonucleotídeos/genética , Filogenia , Síndrome Respiratória Aguda Grave/epidemiologia , Fatores de Tempo
16.
Se Pu ; 22(3): 231-3, 2004 May.
Artigo em Chinês | MEDLINE | ID: mdl-15712904

RESUMO

A new sensitive high performance liquid chromatographic method for the determination of L-cysteine in an enzymatic reaction mixture using ultra violet spectrometric detection was developed. The sample reacted with 5,5'-dithio-bis-nitrobenzoic acid (DTNB) and was analyzed on a Shimadzu VP-ODS column at room temperature, using gradient elution with detection at 330 nm. The L-cysteine chromatographic peak was determined in comparison with derivatives of 2-mercapto ethanol and dithiothreitol. The linear range was 5-950 micromol/L. The recoveries were 99.7%-100.5% and the relative standard deviations (RSDs) were less than 1.3%. The detection limit was 0.8 micromol/L. The method is simple and accurate.


Assuntos
Cromatografia Líquida de Alta Pressão , Cisteína/análise , Cromatografia Líquida de Alta Pressão/métodos , Cisteína/química , Ácido Ditionitrobenzoico/análise , Ditiotreitol/análise , Mercaptoetanol/análise , Espectroscopia Fotoeletrônica/métodos
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