Biomedical subjects

Dong Hua

Publications and source records attributed to Dong Hua.

6 recordsLinked to original sources

PubMed2006

An optimized assay for transcription factor NF-kappaB with dsDNA-coupled microplate.

To develop an EMSA-free assay approach for analyzing the sequence-specific DNA-binding proteins (DBPs), an easy cost-effective dsDNA-coupled plate (dcPlate) was developed in our lab for this purpose. In this paper, the assay conditions of such dcPlate were fully optimized for detecting an important transcription factor, NF-kappaB. The optimized parameters of dcPlate for assay of NF-kappaB were as follows: immobilized DNA probe at the concentration of 25 pmol/100 microL-well, incubation time of 90 min for NF-kappaB binding to dcPlate, primary and secondary antibody concentration of 0.1 microL/100 microL dilution, incubation time of 90 min for primary antibody binding to NF-kappaB, temperature of 25 degrees C for the above process, colorimetric developing time for 30 min. After optimization, the signal was improved three times higher than that from not optimized conditions. The linear colorimetric detection ranges of the purified recombinant NF-kappaB p50 and the cell nuclear extract were from 0.59 to 75 ng/well and 0.313 to 10 microg/well, respectively.

Colorimetry↗

PubMed2006

Quantitative analysis of plasma circulating DNA at diagnosis and during follow-up of breast cancer patients.

The aim of the present project was to quantify the level of plasma DNA in patients with malignant or non-malignant breast diseases and in healthy controls, then investigate whether such measurements have diagnostic or prognostic value. Plasma DNA from 121 women (61 patients with breast cancer, 33 control patients with benign breast diseases and 27 healthy control individuals) was quantified by a real-time quantitative polymerase chain reaction (PCR) method. Plasma DNA concentration in the breast cancer patients (median 65 ng/ml) was significantly higher (P<0.05) than that in the control patients (median 22 ng/ml) or healthy control (median 13 ng/ml). No association was observed between plasma DNA levels and clinicopathological parameters. The data suggest that quantification of plasma circulating DNA may be a valuable complementary diagnostic tool in discriminate breast cancer from unaffected individuals and may be proposed as an early detection test as well as a new, noninvasive assay to follow-up patients.

Breast Neoplasms↗

PubMed2006

Assay of DNA-binding proteins with a dsDNA-coupled plate.

OBJECTIVES: This paper fabricated a cost-effective dsDNA-coupled plate (dcPlate) and applied it to measure the abundance and DNA-binding activity of a DNA-binding protein (DBP). DESIGN AND METHODS: The dcPlate was manufactured by covalently immobilizing an amino-modified oligonucleotide in wells of the plate coated with N-oxysuccinimide esters. The dcPlate was applied to measure the abundance of DNA-binding activity of a DBP in the same four steps, including protein incubation, primary antibody binding, enzyme-linked secondary antibody binding, and colorimetric development. RESULTS: The detections of three purified DBPs including NF-kappaB, AP1 and SP1, and HeLa cell nuclear extract and assays of DNA-binding activity of NF-kappaB p50 to five various DNA sequences demonstrated that dcPlate can be used to measure the abundance of DBPs quantitatively and assay DNA-binding activity of DBPs in high throughputs format. CONCLUSIONS: The homemade cost-effective dcPlate provides a simple and versatile platform for studying DBPs.

Biological Assay↗

PubMed2006

Exonuclease-mediated ELISA-like assay for detecting DNA-binding activity of transcription factors: measurement of activated NF-kappaB.

This paper describes an exonuclease-mediated enzyme-linked immunosorbent assay (ELISA)-like assay (EMEA) for detecting the DNA binding activity of nuclear factor kappaB (NF-kappaB). For EMEA, a special double-stranded DNA (dsDNA)-coupled plate was first prepared by immobilizing a DNA probe on an N-oxysuccinimide ester-coated plate. The immobilized DNA probe, which was internally labeled with digoxigenin (DIG)-dT contained a NF-kappaB binding consensus sequence for capturing activated NF- kappaB in analyzed samples. For measurement, the plate was first incubated with a protein sample and then treated with exonuclease III to eliminate the probes not bound by NF-kappaB. Finally, the probes protected by NF-kappaB were colorimetrically detected by an alkaline phosphatase (AP)-conjugated anti-DIG antibody. The major advantage of EMEA is that it detects NF-kappaB without the need for NF-kappaB antibodies. EMEA may provide a general approach for assays of DNA sequence-specific transcription factors for which specific antibodies are unavailable, expensive, or of insufficient quality.

Base Sequence↗

PubMed2005

Selecting genes by test statistics.

Gene selection is an important issue in analyzing multiclass microarray data. Among many proposed selection methods, the traditional ANOVA F test statistic has been employed to identify informative genes for both class prediction (classification) and discovery problems. However, the F test statistic assumes an equal variance. This assumption may not be realistic for gene expression data. This paper explores other alternative test statistics which can handle heterogeneity of the variances. We study five such test statistics, which include Brown-Forsythe test statistic and Welch test statistic. Their performance is evaluated and compared with that of F statistic over different classification methods applied to publicly available microarray datasets.

Journal Article↗

PubMed2005

Nanoparticle delivery by controlled bacteria.

We present a new approach for nano-object directional delivery by bacteria based upon a taxis-controlled mechanism. In this method, a stimulus is used to direct the bacteria's motion. When carrying nano-objects, the bacteria demonstrated the ability to deliver the "loads" to targets where the stimulus is positioned. The scheme of using taxis for targeted delivery may hold a promising future for a new route to bridge nanotechnology and biotechnology.

Chemotaxis↗