Clinical Assistant Professor, Pathology
Ph.D., The University of Sydney, Biochemistry (1997)
M.D., Shanghai Medical University, Medicine (1987)
Clinical chemistry and therapeutic drug monitoring; Screening, detection and quantification of chromosome translocations in leukemia.
During the development of a testosterone assay by LC-MS/MS, we encountered significant assay interference introduced by blood collection tubes. We examined a number of commonly used blood collection tubes for the presence of interference and its impact on testosterone quantitation.A number of commonly used blood collection tubes were examined by incubation of zero, low and high testosterone concentration samples with them over time, followed by sample preparation using liquid-liquid extraction and analysis by LC-MS/MS. Source of interference was identified by separately incubating blood collection tube coating, stopper and separator gel in clean glass tubes containing zero calibrator.Significant interference was found in some blood collection tubes, with the separator gel identified as the main source. The magnitude of the interference increases over time and mainly affected one of the two testosterone mass transitions used in the quantitation, making it readily detected by the discrepant results obtained by each of the two testosterone mass transitions. We were unable to eliminate the interference by adjustment of the sample preparation procedure, and by changing LC or MS parameters. Accurate quantitation of testosterone is possible when the problematic tubes are avoided, and blood collection tubes free of interference are used instead.Significant LC-MS/MS testosterone assay interference that originated from certain type of blood collection tubes hampered testosterone analysis. Examination of blood collection tube and any other laboratory test tubes for interference should therefore be an integral part of the development and validation of any LC-MS/MS assay used in a clinical diagnostic laboratory.
View details for DOI 10.1016/j.clinbiochem.2012.08.008
View details for Web of Science ID 000312048900035
View details for PubMedID 22971570
To determine whether tubes containing sodium fluoride negatively bias blood glucose concentration by directly comparing glucose concentrations in paired blood samples collected in tubes containing lithium heparin (Li-Heparin) and tubes containing sodium fluoride/potassium oxalate (NaF-KOx).Paired blood samples from a group of patients (n = 1040) were collected in tubes containing Li-Heparin and tubes containing NaF-KOx at the same time. All Li-Heparin samples were centrifuged soon after collection and were kept cool in transport along with NaF-KOx samples, which were centrifuged at the receiving location after an average transport time of 4 h, but immediately before analysis. Glucose concentrations in the paired samples were determined simultaneously by an automated oxidase method.The mean glucose concentrations for NaF-KOx samples and Li-Heparin samples were 5.7 mmol/l and 6.1 mmol/l, respectively, with a mean difference of 0.39 mmol/l.Rapid separation of heparinised blood is superior to fluoride alone for abrogating glycolytic effects on blood glucose measurements in the clinical laboratory.
View details for DOI 10.1136/jcp.2008.062547
View details for Web of Science ID 000268399100017
View details for PubMedID 19638548
Identification of the specific cytogenetic abnormality is one of the critical steps for classification of acute myeloblastic leukemia (AML) which influences the selection of appropriate therapy and provides information about disease prognosis. However at present, the genetic complexity of AML is only partially understood. To obtain a comprehensive, unbiased, quantitative measure, we performed serial analysis of gene expression (SAGE) on CD15(+) myeloid progenitor cells from 22 AML patients who had four of the most common translocations, namely t(8;21), t(15;17), t(9;11), and inv(16). The quantitative data provide clear evidence that the major change in all these translocation-carrying leukemias is a decrease in expression of the majority of transcripts compared with normal CD15(+) cells. From a total of 1,247,535 SAGE tags, we identified 2,604 transcripts whose expression was significantly altered in these leukemias compared with normal myeloid progenitor cells. The gene ontology of the 1,110 transcripts that matched known genes revealed that each translocation had a uniquely altered profile in various functional categories including regulation of transcription, cell cycle, protein synthesis, and apoptosis. Our global analysis of gene expression of common translocations in AML can focus attention on the function of the genes with altered expression for future biological studies as well as highlight genes/pathways for more specifically targeted therapy.
View details for DOI 10.1073/pnas.0509878103
View details for Web of Science ID 000234938300036
View details for PubMedID 16418266
The major challenge to identifying natural sense- antisense (SA) transcripts from public databases is how to determine the correct orientation for an expressed sequence, especially an expressed sequence tag sequence. In this study, we established a set of very stringent criteria to identify the correct orientation of each human transcript. We used these orientation-reliable transcripts to create 26 741 transcription clusters in the human genome. Our analysis shows that 22% (5880) of the human transcription clusters form SA pairs, higher than any previous estimates. Our orientation-specific RT-PCR results along with the comparison of experimental data from previous studies confirm that our SA data set is reliable. This study not only demonstrates that our criteria for the prediction of SA transcripts are efficient, but also provides additional convincing data to support the view that antisense transcription is quite pervasive in the human genome. In-depth analyses show that SA transcripts have some significant differences compared with other types of transcripts, with regard to chromosomal distribution and Gene Ontology-annotated categories of physiological roles, functions and spatial localizations of gene products.
View details for DOI 10.1093/nar/gkh818
View details for Web of Science ID 000224207500011
View details for PubMedID 15356298
Characterization of fusion gene transcripts in leukemia that result from chromosome translocations provides valuable information regarding appropriate treatment and prognosis. However, screening for multiple fusion gene transcripts is difficult with conventional PCR and state-of-the-art real-time PCR and high-density microarrays.We developed a multiplex reverse transcription-PCR (RT-PCR) assay for screening and quantification of fusion gene transcripts in human leukemia cells. Chimeric primers were used that contained gene-specific and universal sequences. PCR amplification of fusion and control gene transcripts was achieved with use of an excess of universal primers to allow the ratio of abundance of fusion gene to endogenous or exogenous controls to be maintained throughout PCR. Multiplex RT-PCR products analyzed by an ABI 310 Genetic Analyzer were consistent with those of duplex RT-PCR (single analytical sample plus control). In addition, multiplex RT-PCR results were analyzed by an assay using an oligonucleotide microarray that contained probes for the splice-junction sequences of various fusion transcripts.The multiplex RT-PCR assay enabled screening of >10 different fusion gene transcripts in a single reaction. RT-PCR followed by analysis with the ABI Prism 310 Genetic Analyzer consistently detected 1 fusion-transcript-carrying leukemia cell in 100-10 000 cells. The assay covered a 1000-fold range. Preliminary results indicate that multiplex RT-PCR products can also be analyzed by hybridization-based microarray assay.The multiplex RT-PCR analyzed by either ABI Prism 310 Genetic Analyzer or microarray provides a sensitive and specific assay for screening of multiple fusion transcripts in leukemia, with the latter an assay that is adaptable to a high-throughput system for clinical screening.
View details for Web of Science ID 000183789600007
View details for PubMedID 12816902
A monoclonal antibody to homovanillic acid (HVA) was prepared by synthesis of a HVA-protein conjugate (HVA-ovalbumin) as an immunogen, immunization of mice, and the subsequent hybridization technique. Monoclonal antibodies were screened on the basis of sensitivity, specificity, and accuracy. An indirect ELISA was developed for quantification of HVA in human urine. The assay was characterized and shown to have high specificity, with cross-reactivities to vanillylmandelic acid and normetanephrine at 0.18% and <0.1%, respectively. The assay coefficients of variation were <10% within the working range of 0.5-40 mg/L. Initial results from testing urine samples of patients with neuroblastoma and other diseases were validated by HPLC, suggesting that this ELISA method is a reliable and convenient system for quantification of HVA in urine and can be used in the mass screening of neuroblastoma in infants.
View details for Web of Science ID 000075221600012
View details for PubMedID 9702954
BACKGROUND and aims. To compare the metabolic effects induced by the anticancer drugs, 6-mercaptopurine (6-MP), 6-thioguanine (6-TG) and 6-methylmercaptopurine riboside (MMPR), which may inhibit the de novo biosynthesis of purine nucleotides or be mis-incorporated into DNA or RNA.Leukaemia cells were grown in culture, exposed to a thiopurine and cell extracts were analyzed for NTPs, dNTPs, drug metabolites and P-Rib-PP.In leukaemia cells, 6-MP was converted to MPR-MP, thio-XMP, thio-GMP, thio-GDP and thio-GTP. Metabolites of 6-TG included thio-XMP, thio-GMP, thio-GDP and thio-GTP, while MMPR-MP was the only major metabolite of MMPR, MMPR (25 microM, 4 h) induced a 16-fold increase in P-Rib-PP and 6-MP (25 microM, 4 h) induced a delayed 5.2-fold increase. MPR-MP, thio-GMP and MMPR-MP are inhibitors of amido phosphoribosyltransferase from leukaemia cells with Ki values of 114 +/- 7.10 microM, 6.20 +/- 2.10 microM and 3.09 +/- 0.30 microM, respectively.The nucleoside-5'-monophosphate derivatives of the 3 thiopurines inhibit amido phosphoribosyltransferase in growing leukaemia cells but there is also an initial inhibition of the further conversion of IMP in the pathway. In growing cells, MMPR acts solely as an inhibitor of de novo purine biosynthesis while 6-TG and to a lesser extent, 6-MP, are converted to significant concentrations of di- and tri-phosphate derivatives which may have other mechanisms of cytotoxicity.
View details for Web of Science ID 000075542700004
View details for PubMedID 9744080