BACKGROUND: Despite the emerging danger of MDR-TB to human beings, there have only been a limited number of drugs developed to treat MDR-TB since 1970. This study investigated the cross-resistance rate between rifampicin (RFP) and rifabutin (RBU) in order to determine the efficacy of rifabutin in treating MDR-TB. In addition, the results of rifabutin were correlated with the rpoB mutations, which are believed to be markers for MDR-TB and RFP resistance. METHODS: The MICs of RBU were tested against 126 clinical isolates of MDR-TB submitted to the clinical laboratory of National Masan TB Hospital in 2004. Five different concentrations (10-160 microgram/ml) were used for the MICs. The detection of the rpoB mutations was performed using a RFP resistance detection kit with a line probe assay(LiPA), which contains the oligonucleotide probes for 5 wide type and 3 specific mutations (513CCA, 516GTC, and 531TTG). The rpoB mutation was determined by direct sequencing. RESULTS: The rate of cross-resistance between RFP and RBU was 70.5%(74/105) at 20 microgram/ml RBU(ed note: How much RFP?) Most mutations (86.3%) occurred in the 524~534 codons. The His526Gln, His526Leu, Leu533Pro, Gln513Glu, and Leu511Pro mutations(Ed note: Is this correct?) were associated with the susceptibilty to RBU. CONCLUSION: Based on the cross-resistance rate between RFP and RBU, RBU may be used effectively in some MDR-TB patients. Therefore, a conventional drug susceptibility test for RBU and a determination of the critical concentration are needed. However, rpoB gene mutation test may be have limited clinical applications in detecting RBU resistance.
Codon ; Humans ; Oligonucleotide Probes ; Rifabutin* ; Rifampin*

No abstract available.
DNA Fingerprinting* ; DNA* ; Oligonucleotide Probes* ; Polymerase Chain Reaction*

OBJECTIVETo establish a highly sensitive fluorometric nanobiosensor for determination of aqueous mercury ions (Hg(2+)) using optimized mercury-specific oligonucleotide (MSO) probes and graphene oxide (GO).

METHODSThe nanobiosensor was assembled by attaching the self-designed MSO(1) (5' end labeled with fluorophore carboxyfluorescein (FAM), denoted as FAM-MSO(1)) and MSO(2) to the surface of GO through strong non-covalent bonding forces. Upon the addition of Hg(2+), the formation of the T-Hg(2+)-T configuration desorbed the FAM-MSO(1) and MSO(2) from the surface of GO, resulting in a restoration of the fluorescence of FAM-MSO(1). Using the specific mispairing of T-Hg(2+)-T and the changes in fluorescent signals in solutions, quantitative analysis of Hg(2+) could be performed.

RESULTSThe average thickness of the prepared GO sheets was only 1.4 nm. For the Hg(2+) nanobiosensor, the optimum concentrations of FAM-MSO(1) and MSO(2) were both 1 µmol/L, the optimum volume of 0.5 g/L GO was 5 µL, and the limit of detection was 10 pmol/L; it had low cross-reactivity with 10 other kinds of non-specific metal ions; the fluorescence recovery efficiency was up to 65% in the re-determination of Hg(2+) after addition of Na(2)S(2)O(3).

CONCLUSIONThe MSO/GO-based nanobiosensor is convenient to operate, highly sensitive, highly specific, highly accurate, and reusable. It can be applied to determine trace amount of Hg(2+) in aqueous solutions.

DNA microarray has become an essential medical genetic diagnostic tool for its high-throughput, miniaturization and automation. The design and selection of oligonucleotide probes are critical for preparing gene chips with high quality. Several sets of probe design software have been developed and are available to perform this work now. Every set of the software aims to different target sequences and shows different advantages and limitations. In this article, the research and development of these sets of software are reviewed in line with three main criteria, including specificity, sensitivity and melting temperature (Tm). In addition, based on the experimental results from literatures, these sets of software are classified according to their applications. This review will be helpful for users to choose an appropriate probe-design software. It will also reduce the costs of microarrays, improve the application efficiency of microarrays, and promote both the research and development (R&D) and commercialization of high-performance probe design software.
Base Sequence ; Oligonucleotide Array Sequence Analysis ; Oligonucleotide Probes ; Sensitivity and Specificity ; Software ; Software Design

OBJECTIVETo investigate HLA genotyping by oligonucleotide arrays.

METHODUnsymmetrical PCR was used to amplify HLA-A gene exon 2, 3. The PCR products were used as templates for hybridization. The oligonucleotide probes were spotted on glass to make microarrays. High signal and specific probes were selected. The effects of the length and location of probes on hybridization signal were studied. A set of computer software was designed for scanning and genotype differentiation.

RESULTThe genotypes of 30 samples analyzed by microarray showed concordance to that by SBT and PCR-SSP.

CONCLUSIONHLA-A genotyping by oligonucleotide array is a good method with advantage of high speed, low cost and high flux.

BACKGROUND: Molecularornucleicacid-based method has been developed for diagnosis as well as epidemiological studies of malaria infection recent years. We developed and evaluated a polymerase chain reaction (PCR)-hybridization assay for its usefulness in diagnosis and genotyping of vivax malaria resurged in South Korea. METHODS: Blood samples were collected from 30 patients diagnosed as vivax malaria and 48 patients with other diseases. The circumsporozoite protein (CSP) gene fragment of Plasmodium vivax was amplified by PCR and hybridized with genotype (VK210 or VK247)-specific oligonucleotide probes. The performance of the assay was evaluated and compared with that by a commercially available immunochromatographic test (ICT; AMRAD, Australia). RESULTS: Twenty-five out of thirty P. vivax-positive blood samples were positive for the PCR-hybridization assay. All products amplified were hybridized only with the VK210-specific probe and showed size polymorphism with approximately 900~ and 865 bp, suggesting of genetic variations of CSP gene. Based on the results of Giemsa-stained blood smear, comparative analysis of test performance demonstrated that sensitivities of the PCR-hybridization assay and ICT were 83.3% and 73.3%, respectively and no false positive results were found. The ktest ratio of two tests yielded results of 0.91 with excellent correlation. CONCLUSOIN: The study suggested that vivax malaria resurged in South Korea has the VK210 genotype of CSP with presence of genetic variants, and that the PCR-hybridization assay is useful for diagnosis as well as genotyping of vivax malaria.
Diagnosis* ; Genetic Variation ; Genotype ; Humans ; Korea ; Malaria ; Malaria, Vivax* ; Oligonucleotide Probes ; Plasmodium vivax ; Polymerase Chain Reaction*

BACKGROUND: Three homemade radiolabeled probes to detect DNA of Mycobacterium tuberculosis by PCR-hybridization (PCRH) assay were compared in order to select the most sensitive and economic probe with the longest lifespan. METHODS: One full length probe, probe 1, prepared by the random priming method and two oligonucleotide probes, probes 2 and 3, prepared by the 5' end-labeling method were designed and assessed for sensitivity, specificity, and life span. The detection limit of each probe was determined on sample membranes containing serially diluted M. tuberculosis DNA from 5 ng to 5 fg on weekly intervals. To assess the specificity of each probe, DNA samples from 4 species of nontuberculous mycobacteria (NTM) and 9 species of bacteria other than mycobacteria were also tested. RESULTS: Each probe with PCRH showed the same detection limits of 50 fg of M. tuberculosis DNA after a 48-hr film exposure time. There were no nonspecific reactions to bacteria when tested for specificity. When we defined the life span of each probe as the longest period for detecting the lowest detection limits of M. tuberculosis DNA, the life spans of probes 1, 2, and 3 after a 3-hour film exposure were 7, 0, and 0 weeks, respectively. For probes 2 and 3, no band was visible even on the day of preparation. The life spans after a 48-hour film exposure were 9, 3, and 2 weeks for probes 1, 2 and 3, respectively. CONCLUSIONS: Probe 1, a full length probe prepared by the random priming method, was more sensitive and was a cheaper probe with a longer life span compared to probes 2 and 3, oligoprobes prepared by the 5' end-labeling method.
Bacteria ; DNA ; Limit of Detection ; Membranes ; Mycobacterium tuberculosis* ; Nontuberculous Mycobacteria ; Oligonucleotide Probes ; Sensitivity and Specificity ; Tuberculosis

The amplification of c-myc oncogene was evaluated in 42 cases of ovarian carcinomas to correlate with clinical parameters. Using oligonucleotide primers, sequences from the c-myc exon-3 gene and from a control gene, tissue plasminogen activator (tPA), were amplified simultaneously by polymerase chain reaction (PCR). After the products of differential PCR (d-PCR) were electrophoresed, slot blot hybridization was performed, and hybridized with P32 dATP-labeled myc and tPA oligonucleotide probes and then autoradiographed. The signal intensities of the two products were quantitated by densitometry and the ratios of two products (c-myc/tPA) were measured. The ovarian carcinomas showed significantly increased amplification of c-myc oncogene Oligonucleoti compared to normal control group (p<0.05). 15 of 42 cases (35.7%) showed various degrees of the MYC gene amplification up to 27 folds in various histologic types of ovarian carcinomas. No significant differences of the MYC gene amplification according to histologic subtypes, tumor action) grades and clinical stages of ovarian carcinomas were present.
Densitometry ; DNA Primers ; Genes, myc ; Oligonucleotide Probes ; Oncogenes* ; Polymerase Chain Reaction* ; Tissue Plasminogen Activator

Polymerase chain reaction (PCR) amplication was used to detect cytomegalovirus (CMV) in tissue culture from the urine of newborns and patients who was suspected CMV infection, Synthetic oligonucleotide primer pairs were used to amplify DNA from the major immediate-early and the phosphoprotein 150 genes of CMV AD 169. Amplified products were detected by gel electrophoresis and by dot-blot hybridization with oligonucleotide probes. We found 12 different tissus culture isolates of CMV after the microimmunoassay using monoclonal antibody to immediate-early antigen. All 12 isolates were positive after PCR amplification. But there was no positive reaction when the same primers and probes were used to amplify herpes simplex virus and human genomic DNA. Twelve urine samples were positive when tested with one or both primer pairs and probes. When compaired tissue culture, detection gel electrophoresis provide a sensitivity of 91% (11/12), dot-blot analysis raised the sensitivity to 100% (12/12). A specificity of both primer was 100%(0/12). We conclude that PCR amplification may be a valuable tool for diagnosing congenital CMV infection.
Cytomegalovirus* ; DNA* ; Electrophoresis ; Humans ; Infant, Newborn ; Oligonucleotide Probes ; Polymerase Chain Reaction* ; Sensitivity and Specificity ; Simplexvirus

BACKGROUND AND OBJECTIVES: This research investigated the association between HLA (human leukocyte antigen) class II alleles and the susceptibility to sudden sensorineural hearing loss, and between HLA class Il alleles and the results of treatment with steroid in the Korean population. MATERIALS AND METHOD: We carried out HLA-DRR1, -DQA1, -DQR1 and -DPRl genotyping in 41 patients with sudden sensorineural hearing loss and in 206 healthy controls. We examined the initial hearing levels at the onset of hearing loss and the final hearing levels after the treatment, and then evaluated for the association with HLA class II alleles. HLA-DRB1, DQA1, BQB1 and DPB1 genotypings were performed by employing the sequence specific oligonucleotide probes (SSOP) method. RESULTS: The frequencies of HLA-DRB1, DQA1, DQB1, and DP31 alleles were not significantly different between the patients and controls. When the association between the results of treatment and HLA alleles was increased, the frequencies of HLA-DRB1X14 (RR= 3.5, p<0.02), -DQA1X03 (RR=4.2, p<0.02) and -DQA1X05 (RR=3.1, p(0.03) significantly increased, but the frequencies of HIA-DQA1X01 (RR=0.2, p<0.004) and -DQB1X06 (RR =0.2, p<0.009) significantly decreased in patients who did not respond to the steroid treatment, compared with the controls. The frequencies of HLA-DQA1X01 (p<0.04) and -DQB1X06 (p<0.02) significantly increased while the frequency of HLA- DQA1X03 (p<0.003) significantly decreased in patients who responded well to steroid, compared with patients who did not. CONCLUSION: These results suggest that the presence of HLA-DRB1X14, DQA1X03 and DQA1X05 might be an useful marker that implys poor prognoses whereas the presence of HLA-DQA101 and DQB1X06 might be a marker implying good prognoses in Korean patients with sudden sensorineural hearing loss.
Alleles* ; Hearing ; Hearing Loss ; Hearing Loss, Sensorineural* ; HLA-DRB1 Chains ; Humans ; Leukocytes ; Oligonucleotide Probes ; Prognosis