Displaying publications 1 - 20 of 1885 in total

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  1. Kathleen, M.M., Samuel, L., Felecia, C., Ng K. H., Lesley, M.B., Kasing, A.
    MyJurnal
    (GTG)5 PCR is a type of repetitive extragenic palindromic (rep)-PCR which amplifies the (GTG)5 repetitive element that lays throughout the bacterial genome. In this study, fifty, thirty-nine and forty-nine unknown bacteria were isolated from aquaculture farms in Miri, Limbang and Lundu, respectively. (GTG)5 PCR was used to screen for clonal diversity among the isolates according to sampling sites. Banding profiles obtained from electrophoresed (GTG)5 PCR products were analyzed by RAPDistance Software to generate a dendrogram of neighbor joining tree (NJT) format. Based on the constructed dendrogram, representative isolates were selected for further identification. Conserved 16S rRNA region of the selected bacteria isolates were amplified and purified DNA products were sequenced. (GTG)5 PCR is useful in differentiation of unknown bacterial isolates and 16S rRNA analysis species identity of the bacteria in Sarawak aquaculture environment. The high diversity of bacteria in aquaculture environment may be caused by contamination from various sources.
    Matched MeSH terms: Polymerase Chain Reaction
  2. Ng ZX, Kuppusamy UR, Tajunisah I, Fong KC, Koay AC, Chua KH
    Br J Ophthalmol, 2012 Feb;96(2):289-92.
    PMID: 22116960 DOI: 10.1136/bjophthalmol-2011-300658
    The receptor for advanced glycation end-products (RAGE) has been implicated in the pathogenesis of diabetic microvascular complications. The aim of this study was to investigate the association between 2245G/A gene polymorphism of the RAGE gene and retinopathy in Malaysian type 2 diabetic patients.
    Matched MeSH terms: Polymerase Chain Reaction
  3. Lou H, Lu Y, Lu D, Fu R, Wang X, Feng Q, et al.
    Am J Hum Genet, 2015 Jul 02;97(1):54-66.
    PMID: 26073780 DOI: 10.1016/j.ajhg.2015.05.005
    Tibetan high-altitude adaptation (HAA) has been studied extensively, and many candidate genes have been reported. Subsequent efforts targeting HAA functional variants, however, have not been that successful (e.g., no functional variant has been suggested for the top candidate HAA gene, EPAS1). With WinXPCNVer, a method developed in this study, we detected in microarray data a Tibetan-enriched deletion (TED) carried by 90% of Tibetans; 50% were homozygous for the deletion, whereas only 3% carried the TED and 0% carried the homozygous deletion in 2,792 worldwide samples (p < 10(-15)). We employed long PCR and Sanger sequencing technologies to determine the exact copy number and breakpoints of the TED in 70 additional Tibetan and 182 diverse samples. The TED had identical boundaries (chr2: 46,694,276-46,697,683; hg19) and was 80 kb downstream of EPAS1. Notably, the TED was in strong linkage disequilibrium (LD; r(2) = 0.8) with EPAS1 variants associated with reduced blood concentrations of hemoglobin. It was also in complete LD with the 5-SNP motif, which was suspected to be introgressed from Denisovans, but the deletion itself was absent from the Denisovan sequence. Correspondingly, we detected that footprints of positive selection for the TED occurred 12,803 (95% confidence interval = 12,075-14,725) years ago. We further whole-genome deep sequenced (>60×) seven Tibetans and verified the TED but failed to identify any other copy-number variations with comparable patterns, giving this TED top priority for further study. We speculate that the specific patterns of the TED resulted from its own functionality in HAA of Tibetans or LD with a functional variant of EPAS1.
    Matched MeSH terms: Polymerase Chain Reaction/methods
  4. Rashid NN, Yusof R, Watson RJ
    Anticancer Res, 2014 Nov;34(11):6557-63.
    PMID: 25368258
    It is well-established that HPV E7 proteins, encoded by human papillomavirus (HPV) genes, frequently associated with cervical cancers bind avidly to the retinoblastoma (RB) family of pocket proteins and disrupt their association with members of the E2F transcription factor family. Our previous study showed that the repressive p130-dimerization partner, RB-like, E2F and multi-vulval class (DREAM) complex was disrupted by HPV16 E7 proteins in order to maintain the viral replication in CaSki cells. However, we would like to address whether the activator B-myb-DREAM complex is critical in regulating the replication and mitosis phase since our previous study showed increased B-myb-DREAM expression in HPV-transformed cell lines when compared to control cells.
    Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction; Real-Time Polymerase Chain Reaction
  5. Sideek MA, Smith J, Menz C, Adams JRJ, Cowin AJ, Gibson MA
    Int J Mol Sci, 2017 Oct 09;18(10).
    PMID: 28991210 DOI: 10.3390/ijms18102114
    Latent transforming growth factor-β-1 binding protein-2 (LTBP-2) belongs to the LTBP-fibrillin superfamily of extracellular proteins. Unlike other LTBPs, LTBP-2 does not covalently bind transforming growth factor-β1 (TGF-β1) but appears to be implicated in the regulation of TGF-β1 bioactivity, although the mechanisms are largely unknown. In experiments originally designed to study the displacement of latent TGF-β1 complexes from matrix storage, we found that the addition of exogenous LTBP-2 to cultured human MSU-1.1 fibroblasts caused an increase in TGF-β1 levels in the medium. However, the TGF-β1 increase was due to an upregulation of TGF-β1 expression and secretion rather than a displacement of matrix-stored TGF-β1. The secreted TGF-β1 was mainly in an inactive form, and its concentration peaked around 15 h after addition of LTBP-2. Using a series of recombinant LTBP-2 fragments, the bioactivity was identified to a small region of LTBP-2 consisting of an 8-Cys motif flanked by four epidermal growth factor (EGF)-like repeats. The LTBP-2 stimulation of TGF-β expression involved the phosphorylation of both Akt and p38 mitogen-activated protein kinase (MAPK) signalling proteins, and specific inactivation of each protein individually blocked TGF-β1 increase. The search for the cell surface receptor mediating this LTBP-2 activity proved inconclusive. Inhibitory antibodies to integrins β1 and αVβ5 showed no reduction of LTBP-2 stimulation of TGF-β1. However, TGF-β1 upregulation was partially inhibited by anti-αVβ3 integrin antibodies, suggestive of a direct or indirect role for this integrin. Overall, the study indicates that LTBP-2 can directly upregulate cellular TGF-β1 expression and secretion by interaction with cells via a short central bioactive region. This may be significant in connective tissue disorders involving aberrant TGF-β1 signalling.
    Matched MeSH terms: Polymerase Chain Reaction
  6. Yusof, R., Abdul Rahman, P.S., Rahim, Z.H.A.
    Ann Dent, 1999;6(1):-.
    MyJurnal
    The application of PCR technique in genetic screening was demonstrated using the genetic materials from buccal cells of the students in the class. Two factors were taken into consideration when designing the experiments. The DNA region to be amplified should not be associated with any disease state. This is to eliminate any emotional and ethical problems associated with the experiments. In this practical, the presence and absence of a 38 bp sequence in the intron of COLIA2 gene were studied. The students were also shown on how to analyse the presence of homozygous and heterozygous alleles and the genetic variations that might be observed in the different ethnic groups of students. Another factor was the time taken to complete the experiment. Our experience showed that this experiment would take at least six hours to obtain and analyse the results. It is therefore suitable to be used in class teaching.
    Matched MeSH terms: Polymerase Chain Reaction
  7. Low JS, Chin YM, Mushiroda T, Kubo M, Govindasamy GK, Pua KC, et al.
    PLoS One, 2016;11(1):e0145774.
    PMID: 26730743 DOI: 10.1371/journal.pone.0145774
    BACKGROUND: Nasopharyngeal carcinoma (NPC) is a neoplasm of the epithelial lining of the nasopharynx. Despite various reports linking genomic variants to NPC predisposition, very few reports were done on copy number variations (CNV). CNV is an inherent structural variation that has been found to be involved in cancer predisposition.

    METHODS: A discovery cohort of Malaysian Chinese descent (NPC patients, n = 140; Healthy controls, n = 256) were genotyped using Illumina® HumanOmniExpress BeadChip. PennCNV and cnvPartition calling algorithms were applied for CNV calling. Taqman CNV assays and digital PCR were used to validate CNV calls and replicate candidate copy number variant region (CNVR) associations in a follow-up Malaysian Chinese (NPC cases, n = 465; and Healthy controls, n = 677) and Malay cohort (NPC cases, n = 114; Healthy controls, n = 124).

    RESULTS: Six putative CNVRs overlapping GRM5, MICA/HCP5/HCG26, LILRB3/LILRA6, DPY19L2, RNase3/RNase2 and GOLPH3 genes were jointly identified by PennCNV and cnvPartition. CNVs overlapping GRM5 and MICA/HCP5/HCG26 were subjected to further validation by Taqman CNV assays and digital PCR. Combined analysis in Malaysian Chinese cohort revealed a strong association at CNVR on chromosome 11q14.3 (Pcombined = 1.54x10-5; odds ratio (OR) = 7.27; 95% CI = 2.96-17.88) overlapping GRM5 and a suggestive association at CNVR on chromosome 6p21.3 (Pcombined = 1.29x10-3; OR = 4.21; 95% CI = 1.75-10.11) overlapping MICA/HCP5/HCG26 genes.

    CONCLUSION: Our results demonstrated the association of CNVs towards NPC susceptibility, implicating a possible role of CNVs in NPC development.

    Matched MeSH terms: Polymerase Chain Reaction
  8. Hafidh RR, Hussein SZ, MalAllah MQ, Abdulamir AS, Abu Bakar F
    Curr Cancer Drug Targets, 2018;18(8):807-815.
    PMID: 29141549 DOI: 10.2174/1568009617666171114144236
    BACKGROUND: Citrus bioactive compounds, as active anticancer agents, have been under focus by several studies worldwide. However, the underlying genes responsible for the anticancer potential have not been sufficiently highlighted.

    OBJECTIVES: The current study investigated the gene expression profile of hepatocellular carcinoma, HepG2, cells after treatment with Limonene.

    METHODS: The concentration that killed 50% of HepG2 cells was used to elucidate the genetic mechanisms of limonene anticancer activity. The apoptotic induction was detected by flow cytometry and confocal fluorescence microscope. Two of the pro-apoptotic events, caspase-3 activation and phosphatidylserine translocation were manifested by confocal fluorescence microscopy. Highthroughput real-time PCR was used to profile 1023 cancer-related genes in 16 different gene families related to the cancer development.

    RESULTS: In comparison to untreated cells, limonene increased the percentage of apoptotic cells up to 89.61%, by flow cytometry, and 48.2% by fluorescence microscopy. There was a significant limonene- driven differential gene expression of HepG2 cells in 15 different gene families. Limonene was shown to significantly (>2log) up-regulate and down-regulate 14 and 59 genes, respectively. The affected gene families, from the most to the least affected, were apoptosis induction, signal transduction, cancer genes augmentation, alteration in kinases expression, inflammation, DNA damage repair, and cell cycle proteins.

    CONCLUSION: The current study reveals that limonene could be a promising, cheap, and effective anticancer compound. The broad spectrum of limonene anticancer activity is interesting for anticancer drug development. Further research is needed to confirm the current findings and to examine the anticancer potential of limonene along with underlying mechanisms on different cell lines.

    Matched MeSH terms: Real-Time Polymerase Chain Reaction
  9. Zahari Z, Salleh MR, Zahri Johari MK, Musa N, Ismail R
    Malays J Med Sci, 2011 Oct;18(4):44-57.
    PMID: 22589672 MyJurnal
    The dopamine D2 receptor gene (DRD2) plays a role in many diseases such as schizophrenia, Parkinson's disease, and addictive behaviour. Methods currently available for the detection of DRD2 polymorphisms are costly and cannot detect all 8 polymorphisms of our research interest simultaneously (Val96Ala, Leu141Leu, Val154Ile, Pro310Ser, Ser311Cys, TaqI A, A-241G, and -141C Ins/Del). Therefore, we developed a nested multiplex polymerase chain reaction (PCR) for simultaneous detection of these polymorphisms.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  10. Wong MY, Smart CD
    Plant Dis, 2012 Sep;96(9):1365-1371.
    PMID: 30727148 DOI: 10.1094/PDIS-07-11-0593-SR
    A DNA macroarray was previously developed to detect major fungal and oomycete pathogens of solanaceous crops. To provide a convenient alternative for researchers with no access to X-ray film-developing facilities, specific CCD cameras or Chemidoc XRS systems, a chromogenic detection method with sensitivity comparable with chemiluminescent detection, has been developed. A fungal (Stemphylium solani) and an oomycete (Phytophthora capsici) pathogen were used to develop the protocol using digoxigenin (DIG)-labeled targets. The internal transcribed spacer (ITS) region of the nuclear ribosomal DNA (rDNA), including ITS1, 5.8S rDNA, and ITS2, was used as the target gene and polymerase chain reaction amplified as in the previous protocol. Various amounts of species-specific oligonucleotides on the array, quantities of DIG-labeled ITS amplicon, and hybridization temperatures were tested. The optimal conditions for hybridization were 55°C for 2 h using at least 10 pmol of each species-specific oligonucleotide and labeled target at 10 ng/ml of hybridization buffer. Incubation of the hybridized array with anti-DIG conjugated alkaline phosphatase substrates, NBT/BCIP, produced visible target signals between 1 and 3 h compared with 1 h in chemiluminescent detection. Samples from pure cultures, soil, and artificially inoculated plants were also used to compare the detection using chemiluminescent and chromogenic methods. Chromogenic detection was shown to yield similar results compared with chemiluminescent detection in regard to signal specificity, duration of hybridization between the array and targets, and cost, though it takes 1 to 2 h longer for the visualization process, thus providing a convenient alternative for researchers who lack darkroom facilities. To our knowledge, this is the first report of DNA macroarray detection of plant pathogens using a chromogenic method.
    Matched MeSH terms: Polymerase Chain Reaction
  11. Phua AC, Abdullah RB, Mohamed Z
    J. Reprod. Dev., 2003 Aug;49(4):307-11.
    PMID: 14967923
    Sex determination of livestock is performed to achieve the objectives of livestock breeding programmes. Techniques for sex determination have evolved from karyotyping to detecting Y-specific antigens and recently to the polymerase chain reaction (PCR), which appears to be the most sensitive, accurate, rapid and reliable method to date. In this study, a PCR-based sex determination method for potential application in goat breeding programmes was developed. Primers were designed to amplify a portion of the X amelogenin gene (Aml-X) on the X chromosome to give a 300 bp product and Sry gene on the Y chromosome to give a 116 bp product. PCR optimization was performed using DNA template extracted from a whole blood sample of Jermasia goats (German Fawn x Katjang) of both sexes. It was possible to identify the sex chromosomes by amplifying both male- and female-specific genes simultaneously in a duplex reaction with males yielding two bands and females yielding one band. The Aml-X primer set, which served as an internal control primer, did not interfere with amplification of the Y-specific sequence even when a low amount of DNA (1 ng) was used. The duplex reaction subjected to a blind test showed 100% (14/14) concordance, proving its accuracy and reliability. The primer sets used were found to be highly specific and were suitable for gender selection of goats.
    Matched MeSH terms: Polymerase Chain Reaction/methods*
  12. Xiu L, Binder RA, Alarja NA, Kochek K, Coleman KK, Than ST, et al.
    J Clin Virol, 2020 07;128:104391.
    PMID: 32403008 DOI: 10.1016/j.jcv.2020.104391
    BACKGROUND: During the past two decades, three novel coronaviruses (CoVs) have emerged to cause international human epidemics with severe morbidity. CoVs have also emerged to cause severe epidemics in animals. A better understanding of the natural hosts and genetic diversity of CoVs are needed to help mitigate these threats.

    OBJECTIVE: To design and evaluate a molecular diagnostic tool for detection and identification of all currently recognized and potentially future emergent CoVs from the Orthocoronavirinae subfamily.

    STUDY DESIGN AND RESULTS: We designed a semi-nested, reverse transcription RT-PCR assay based upon 38 published genome sequences of human and animal CoVs. We evaluated this assay with 14 human and animal CoVs and 11 other non-CoV respiratory viruses. Through sequencing the assay's target amplicon, the assay correctly identified each of the CoVs; no cross-reactivity with 11 common respiratory viruses was observed. The limits of detection ranged from 4 to 4 × 102 copies/reaction, depending on the CoV species tested. To assess the assay's clinical performance, we tested a large panel of previously studied specimens: 192 human respiratory specimens from pneumonia patients, 5 clinical specimens from COVID-19 patients, 81 poultry oral secretion specimens, 109 pig slurry specimens, and 31 aerosol samples from a live bird market. The amplicons of all RT-PCR-positive samples were confirmed by Sanger sequencing. Our assay performed well with all tested specimens across all sample types.

    CONCLUSIONS: This assay can be used for detection and identification of all previously recognized CoVs, including SARS-CoV-2, and potentially any emergent CoVs in the Orthocoronavirinae subfamily.

    Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction/methods*
  13. Patel K, Klena J, Lo MK
    Methods Mol Biol, 2023;2682:25-31.
    PMID: 37610571 DOI: 10.1007/978-1-0716-3283-3_2
    From its discovery in Malaysia in the late 1990s, the spillover of the Nipah virus from its pteropid reservoir into the human population has resulted in sporadic outbreaks of fatal encephalitis and respiratory disease. In this chapter, we revise a previously described quantitative reverse transcription polymerase chain reaction method, which now utilizes degenerate nucleotides at certain positions in the probe and the reverse primer to accommodate the sequence heterogeneity observed within the Nipah henipavirus species.
    Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
  14. Divis PC, Shokoples SE, Singh B, Yanow SK
    Malar J, 2010 Nov 30;9:344.
    PMID: 21114872 DOI: 10.1186/1475-2875-9-344
    BACKGROUND: The misdiagnosis of Plasmodium knowlesi by microscopy has prompted a re-evaluation of the geographic distribution, prevalence and pathogenesis of this species using molecular diagnostic tools. In this report, a specific probe for P. knowlesi, that can be used in a previously described TaqMan real-time PCR assay for detection of Plasmodium spp., and Plasmodium falciparum, Plasmodium vivax, Plasmodium malariae and Plasmodium ovale, was designed and validated against clinical samples.

    METHODS: A hydrolysis probe for a real-time PCR assay was designed to recognize a specific DNA sequence within the P. knowlesi small subunit ribosomal RNA gene. The sensitivity, linearity and specificity of the assay were determined using plasmids containing P. knowlesi DNA and genomic DNA of P. falciparum, P. knowlesi, P. malariae, P. ovale and P. vivax isolated from clinical samples. DNA samples of the simian malaria parasites Plasmodium cynomolgi and Plasmodium inui that can infect humans under experimental conditions were also examined together with human DNA samples.

    RESULTS: Analytical sensitivity of the P. knowlesi-specific assay was 10 copies/μL and quantitation was linear over a range of 10-106 copies. The sensitivity of the assay is equivalent to nested PCR and P. knowlesi DNA was detected from all 40 clinical P. knowlesi specimens, including one from a patient with a parasitaemia of three parasites/μL of blood. No cross-reactivity was observed with 67 Plasmodium DNA samples (31 P. falciparum, 23 P. vivax, six P. ovale, three P. malariae, one P. malariae/P. ovale, one P. falciparum/P. malariae, one P. inui and one P. cynomolgi) and four samples of human DNA.

    CONCLUSIONS: This test demonstrated excellent sensitivity and specificity, and adds P. knowlesi to the repertoire of Plasmodium targets for the clinical diagnosis of malaria by real-time PCR assays. Furthermore, quantitation of DNA copy number provides a useful advantage over other molecular assays to investigate the correlation between levels of infection and the spectrum of disease.

    Matched MeSH terms: Polymerase Chain Reaction/methods*
  15. Khazani NA, Noor NZ, Yean Yean C, Hasan H, Suraiya S, Mohamad S
    J Trop Med, 2017;2017:7210849.
    PMID: 28386286 DOI: 10.1155/2017/7210849
    Klebsiella pneumoniae and Haemophilus influenzae are two common pathogens associated with respiratory tract infections. The identification of these pathogens using conventional molecular diagnostic tests requires trained personnel, cold-chain transportation, and storage-dependance, which does not render them user-friendly. The aim of this study was to develop a thermostabilized, cold-chain-free, one-step multiplex PCR for simultaneous detection of K. pneumoniae and H. influenzae. The multiplex PCR assay was designed to amplify the php gene of K. pneumoniae (202 bp) and p6 gene of H. influenzae (582 bp). In addition, the specific primer to amplify glm gene of Helicobacter pylori (105 bp) was included as an internal amplification control. Subsequently, the designed primers and all PCR reagents were thermostabilized by lyophilization. The stability of the thermostabilized PCR was evaluated using the Q(10) method. The sensitivity and specificity of performances for thermostabilized PCR were evaluated using 127 clinical isolates and were found to be 100% sensitive and specific. The thermostabilized PCR mix was found to be stable for 30 days and the Q10 accelerated stability was found to be 3.02 months. A cold-chain-free, PCR assay for easy, rapid, and simultaneous detection of K. pneumoniae and H. influenzae was successfully developed in this study.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction
  16. Ponnampalam SN, Kamaluddin NR, Zakaria Z, Matheneswaran V, Ganesan D, Haspani MS, et al.
    Oncol Rep, 2017 Jan;37(1):10-22.
    PMID: 28004117 DOI: 10.3892/or.2016.5285
    The aims of the present study were to undertake gene expression profiling of the blood of glioma patients to determine key genetic components of signaling pathways and to develop a panel of genes that could be used as a potential blood-based biomarker to differentiate between high and low grade gliomas, non-gliomas and control samples. In this study, blood samples were obtained from glioma patients, non-glioma and control subjects. Ten samples each were obtained from patients with high and low grade tumours, respectively, ten samples from non-glioma patients and twenty samples from control subjects. Total RNA was isolated from each sample after which first and second strand synthesis was performed. The resulting cRNA was then hybridized with the Agilent Whole Human Genome (4x44K) microarray chip according to the manufacturer's instructions. Universal Human Reference RNA and samples were labeled with Cy3 CTP and Cy5 CTP, respectively. Microarray data were analyzed by the Agilent Gene Spring 12.1V software using stringent criteria which included at least a 2-fold difference in gene expression between samples. Statistical analysis was performed using the unpaired Student's t-test with a p<0.01. Pathway enrichment was also performed, with key genes selected for validation using droplet digital polymerase chain reaction (ddPCR). The gene expression profiling indicated that were a substantial number of genes that were differentially expressed with more than a 2-fold change (p<0.01) between each of the four different conditions. We selected key genes within significant pathways that were analyzed through pathway enrichment. These key genes included regulators of cell proliferation, transcription factors, cytokines and tumour suppressor genes. In the present study, we showed that key genes involved in significant and well established pathways, could possibly be used as a potential blood-based biomarker to differentiate between high and low grade gliomas, non-gliomas and control samples.
    Matched MeSH terms: Polymerase Chain Reaction/methods
  17. Shanmugam H, Eow GI, Nadarajan VS
    Malays J Pathol, 2009 Jun;31(1):63-6.
    PMID: 19694316 MyJurnal
    Adult T-cell leukaemia/lymphoma (ATLL) is a rare T lymphoproliferative disorder which is aetiologically linked with human T-cell lymphotropic virus type-1 (HTLV-1). HTLV-1 is endemic in Japan, Caribbean and Africa. The highest incidence of ATLL is in Japan although sporadic cases have been reported elsewhere in the world. We describe a case of ATLL with an unusual presentation which we believe is the first reported case of ATLL in Malaysia based on our literature search. A 51-year-old Indian lady was referred to University Malaya Medical Centre for an incidental finding of lymphocytosis while being investigated for pallor and giddiness. Clinical examination revealed bilateral shotty cervical lymph nodes with no hepato-splenomegaly or skin lesions. Laboratory investigations showed absolute lymphocytosis (38 x 10(9)/L) with a mildly increased serum lactate dehydrogenase. The peripheral blood smear showed the presence of predominantly small to medium sized, non-flower lymphocytes. The bone marrow showed similar findings of prominent lymphocytosis. Immunophenotyping of the bone marrow mononuclear cells showed CD3+, CD4+, CD5+, CD7- and CD25+ which is characteristic of ATLL phenotype. HTLV-1 infection was confirmed by the presence of HTLV-1 proviral DNA in the tumor cells using conventional Polymerase Chain Reaction (PCR) and real-time PCR. Here, we discuss the pathogenesis and characteristics of ATLL as well as the detection of HTLV-1 by real time PCR.
    Matched MeSH terms: Polymerase Chain Reaction
  18. Yip KT, Das PK, Suria D, Lim CR, Ng GH, Liew CC
    J Exp Clin Cancer Res, 2010;29:128.
    PMID: 20846378 DOI: 10.1186/1756-9966-29-128
    BACKGROUND: Colorectal cancer (CRC) screening is key to CRC prevention and mortality reduction, but patient compliance with CRC screening is low. We previously reported a blood-based test for CRC that utilizes a seven-gene panel of biomarkers. The test is currently utilized clinically in North America for CRC risk stratification in the average-risk North American population in order to improve screening compliance and to enhance clinical decision making.
    METHODS: In this study, conducted in Malaysia, we evaluated the seven-gene biomarker panel validated in a North American population using blood samples collected from local patients. The panel employs quantitative RT-PCR (qRT-PCR) to analyze gene expression of the seven biomarkers (ANXA3, CLEC4D, TNFAIP6, LMNB1, PRRG4, VNN1 and IL2RB) that are differentially expressed in CRC patients as compared with controls. Blood samples from 210 patients (99 CRC and 111 controls) were collected, and total blood RNA was isolated and subjected to quantitative RT-PCR and data analysis.
    RESULTS: The logistic regression analysis of seven-gene panel has an area under the curve (AUC) of 0.76 (95% confidence interval: 0.70 to 0.82), 77% specificity, 61% sensitivity and 70% accuracy, comparable to the data obtained from the North American investigation of the same biomarker panel.
    CONCLUSIONS: Our results independently confirm the results of the study conducted in North America and demonstrate the ability of the seven biomarker panel to discriminate CRC from controls in blood samples drawn from a Malaysian population.
    Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
  19. Yamada M, Shishito N, Nozawa Y, Uni S, Nishioka K, Nakaya T
    Trop Med Health, 2017;45:26.
    PMID: 29118653 DOI: 10.1186/s41182-017-0067-4
    Background: Dirofilaria ursi is a filarial nematode that parasitizes the subcutaneous tissues of the American black bear (Ursus americanus) and Japanese black bear (Ursus thiabetanus japonicus). D. ursi that has parasitized black bears has the potential to subsequently infect humans. In addition, extra-gastrointestinal anisakiasis is less common in Japan.

    Case presentation: We report a case of ventral subcutaneous anisakiasis and dorsal subcutaneous dirofilariasis that was acquired in Fukushima, in the northern part of Japan. The patient was an 83-year-old Japanese female, and subcutaneous parasitic granulomas were present on her left abdomen (near the navel) and left scapula. A pathological examination of the surgically dissected tissue sections from each region demonstrated eosinophilic granulomas containing different species of parasites. To enable the morphological and molecular identification of these parasites, DNA was extracted from paraffin-embedded sections using DEXPAT reagent, and the cytochrome oxidase 2 (COX2), internal transcribed spacer 1 (ITS1), 5.8S and ITS2 regions of the Anisakis larvae, and the 5S rRNA region of the male Dirofilaria were sequenced. The PCR products were examined and compared with DNA databases. Molecular analysis of the COX2 and 5S rRNA sequences of each worm revealed that the nematode found in the ventral region belonged to Anisakis simplex sensu stricto (s.s.) and the male Dirofilaria found in the dorsal region was classified as D. ursi.

    Conclusion: The present case showed a combined human case of D. ursi and A. simplex s.s. infections in subcutaneous tissues. The results of this study will contribute to the identification of unknown parasites in histological sections.
    Matched MeSH terms: Polymerase Chain Reaction
  20. Chua KB, Mustafa B, Abdul Wahab AH, Chem YK, Khairul AH, Kumarasamy V, et al.
    Malays J Pathol, 2011 Jun;33(1):13-20.
    PMID: 21874746
    A prospective study was carried out to evaluate the sensitivity of dengue NS1 antigen-capture ELISA in comparison with dengue virus isolation, conventional RT-PCR and real-time RT-PCR for laboratory confirmation of acute dengue based on single-acute serum samples. Four primary healthcare centres were involved to recruit patients with clinical diagnosis of dengue illness. Patient's demographic, epidemiological and clinical information were collected on a standardized data entry form and 5 ml of venous blood was collected upon consent. In the laboratory, six types of laboratory tests were performed on each of the collected acute serum sample. Of the 558 acute serum samples collected from 558 patients with clinical diagnosis of dengue from mid-August 2006 to March 2009, 174 serum samples were tested positive by the dengue NS1 antigen-capture ELISA, 77 by virus isolation, 92 by RT-PCR and 112 by real-time RT-PCR. A total of 190 serum samples were tested positive by either one or a combination of the four methods whereas, only 59 serum samples were tested positive by all four methods. Thus, based on single-acute serum samples, 190 of the 558 patients (34.1%) were laboratory-confirmed acute dengue. The overall test sensitivity was 91.6%, 40.5%, 48.4% and 58.9% for dengue NS1 antigen-capture ELISA, virus isolation, conventional RT-PCR and real-time RT-PCR respectively. Statistically, dengue NS1 antigen-capture ELISA was the most sensitive and virus isolation was the least sensitive test for the laboratory confirmation of acute dengue based on single-acute serum specimens. Real-time RT-PCR was significantly more sensitive than the conventional RT-PCR.
    Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction/methods*
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