Biomedical subjects

Ger van den Engh

Publications and source records attributed to Ger van den Engh.

4 recordsLinked to original sources

PubMed2003

High-speed cell sorting: fundamentals and recent advances.

Cell sorters have undergone dramatic technological improvements in recent years. Driven by the increased ability to differentiate between cell types, modern advances have yielded a new generation of cytometers, known as high-speed cell sorters. These instruments are capable of higher throughput than traditional sorters and can distinguish subtler differences between particles by measuring and processing more optical parameters in parallel. These advances have expanded their use to facilitate genomic and proteomic discovery, and as vehicles for many emerging cell-based therapies. High-speed cell sorting is becoming established as an essential research tool across a broad range of scientific fields and is poised to play a pivotal role in the latest therapeutic modalities.

Cell Separation↗

PubMed2002

Use of fluorescent sequence-specific polyamides to discriminate human chromosomes by microscopy and flow cytometry.

In this paper, we demonstrate the use of synthetic polyamide probes to fluorescently label heterochromatic regions on human chromosomes for discrimination in cytogenetic preparations and by flow cytometry. Polyamides bind to the minor groove of DNA in a sequence-specific manner. Unlike conventional sequence-specific DNA or RNA probes, polyamides can recognize their target sequence without the need to subject chromosomes to harsh denaturing conditions. For this study, we designed and synthesized a polyamide to target the TTCCA-motif repeated in the heterochromatic regions of chromosome 9, Y and 1. We demonstrate that the fluorescently labeled polyamide binds to its target sequence in both conventional cytogenetic preparations of metaphase chromosomes and suspended chromosomes without denaturation. Chromosomes 9 and Y can be discriminated and purified by flow sorting on the basis of polyamide binding and Hoechst 33258 staining. We generate chromosome 9- and Y-specific 'paints' from the sorted fractions. We demonstrate the utility of this technology by characterizing the sequence of an olfactory receptor gene that is duplicated on multiple chromosomes. By separating chromosome 9 from chromosomes 10-12 on the basis of polyamide fluorescence, we determine and differentiate the haplotypes of the highly similar copies of this gene on chromosomes 9 and 11.

Base Sequence↗

PubMed2002

Human prostate epithelial cell-type cDNA libraries and prostate expression patterns.

BACKGROUND: Transcriptome analysis is a powerful approach to uncovering genes responsible for diseases such as prostate cancer. Ideally, one would like to compare the transcriptomes of a cancer cell and its normal counterpart for differences. METHODS: Prostate luminal and basal epithelial cell types were isolated and cell-type-specific cDNA libraries were constructed. Sequence analysis of cDNA clones generated 505 luminal cell genes and 560 basal cell genes. These sequences were deposited in a public database for expression analysis. RESULTS: From these sequences, 119 unique luminal expressed sequence tags (ESTs) were extracted and assembled into a luminal-cell transcriptome set, while 154 basal ESTs were extracted and assembled into a basal-cell set. Interlibrary comparison was performed to determine representation of these sequences in cDNA libraries constructed from prostate tumors, PIN, cell lines. CONCLUSIONS: Our analysis showed that a significant number of epithelial cell genes were not represented in the various transcriptomes of prostate tissues, suggesting that they might be underrepresented in libraries generated from tissue containing multiple cell types. Although both luminal and basal cell types are epithelial, their transcriptomes are more divergent from each other than expected, underscoring their functional difference (secretory vs. nonsecretory). Tumor tissues show different expression of luminal and basal genes, with perhaps a trend towards expression of basal genes in advanced diseases.

DNA, Complementary↗

PubMed2002

Trapping of DNA by dielectrophoresis.

Under suitable conditions, a DNA molecule in solution will develop a strong electric dipole moment. This induced dipole allows the molecule to be manipulated with field gradients, in a phenomenon known as dielectrophoresis (DEP). Pure dielectrophoretic motion of DNA requires alternate current (AC) electric fields to suppress the electrophoretic effect of the molecules net charge. In this paper, we present two methods for measuring the efficiency of DEP for trapping DNA molecules as well as a set of quantitative measurements of the effects of strand length, buffer composition, and frequency of the applied electric field. A simple configuration of electrodes in combination with a microfluidic flow chamber is shown to increase the concentration of DNA in solution by at least 60-fold. These results should prove useful in designing practical microfluidic devices employing this phenomenon either for separation or concentration of DNA.

Bacteriophage lambda↗