Nanoparticle-based energy transfer for rapid and simple detection of protein glycosylation.
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Energy Transfer↗
Biomedical subjects
Dohoon Lee
Publications and source records attributed to Dohoon Lee.
Gold nanoparticle-enhanced secondary ion mass spectrometry imaging of peptides on self-assembled monolayers.
We demonstrate the use of gold nanoparticles (AuNPs) to enhance the secondary ion emission of peptides in time-of-flight secondary ion mass spectrometry (TOF-SIMS). The signal intensity of peptides adsorbed onto AuNPs was significantly increased when compared to that of self-assembled monolayers (SAMs). This gold nanoparticle-enhanced SIMS, termed NE-SIMS, enabled the sensitive detection of subtle modifications of peptides, such as phosphorylation. From a quantitative analysis of the amounts of adsorbed peptides and AuNPs on SAMs using quartz crystal microbalance and surface plasmon resonance spectroscopy, the ratio of peptide molecule to AuNP on amine-SAMs was revealed to be 18-19:1. When considering the ratio of peptide to matrix (1:10(3)-10(6)) employed in a matrix-enhanced SIMS, the use of AuNPs gave rise to a significantly increased secondary ion emission of peptides. Peptides were adsorbed onto patterned AuNPs on SAMs using a microfluidic system, and well-contrasted molecular ion images were obtained. NE-SIMS is expected to be applied to a chip-based analysis of modification of biomolecules in a label-free manner.
Gold↗
Expression profiling of proteins in L-threonine biosynthetic pathway of Escherichia coli by using antibody microarray.
We demonstrate the use of an antibody (Ab) microarray for a comparative expression profiling of proteins in an L-threonine biosynthetic pathway of Escherichia coli between a parental strain (W3110) and L-threonine overproducing mutant (TF5015). On the basis of a global comparative transcriptome analysis between the two strains, 28 analytical target proteins were selected and subjected to a production of polyclonal Abs against them. An Ab microarray was constructed by spotting a set of produced antibodies on a glass slide, and was employed for a comparative expression profiling of the proteins between the two strains by a two-color fluorescence assay method. The performance of the Ab microarray was evaluated with respect to cross-reactivity of the antibodies, dye-labeling efficiency, and the nature of antigenic proteins. Of these, the cross-reactivity of the used antibodies was found to mainly cause the deviation of the observed expression ratios from the expected ones. To offset the deviations, correction factors were derived from a statistical analysis and introduced. As a result, ten proteins were categorized to be up-regulated, while one was down-regulated in TF5015. Expression profiling of proteins using the Ab microarray was further verified by comparison with Western blotting and 2-DE.
Antibodies, Bacterial↗
Making it in America: high school completion by immigrant and native youth.
Using data from the National Longitudinal Study of Adolescent Health (Add Health), we find that first-generation youth of Hispanic, Asian, and African heritage obtain more education than their parents, but the second generation and third or higher generations lose ground. Differences in dropout rates by race-ethnicity and immigrant generation are driven by differences in human, cultural, and social capital. Low levels of family human capital, school social capital, and community social capital place the children of immigrants at risk of dropping out. However, cultural capital and immigrant optimism buffer first-generation Hispanic youth and the children of Asian immigrants from the risk of dropping out of high school. While human and social capital resources improve with immigrant generation, cultural capital diminishes.
Acculturation↗
Preparation of a magnetically switchable bio-electrocatalytic system employing cross-linked enzyme aggregates in magnetic mesocellular carbon foam.
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Carbon↗
Kinetic and equilibrium binding analysis of protein-ligand interactions at poly(amidoamine) dendrimer monolayers.
The interaction of streptavidin (SA) with a biotinylated surface has been of great interest in the development of an interfacial layer for protein immobilization based on self-assembled monolayers (SAMs) and polymeric layers. Here, we demonstrate the unique characteristics of protein-ligand interactions on dendrimer monolayers based on kinetic and equilibrium binding analyses. With amine-ended poly(amidoamine) dendrimers from the first (G1) to fourth (G4) generation, the formation of even, compact dendrimer monolayers on gold was confirmed using FT-IR spectroscopy and ellipsometry. For the SA-biotin interaction, quantitative analysis of bound SA using surface plasmon resonance showed that the saturation binding level of SA was fairly higher in all dendrimer layers when compared to other tested systems of 11-mercaptoundecylamine SAMs and a poly(L-lysine) layer. Kinetic studies revealed that the initial binding rate of SA up to the saturation level was 2-fold higher in all dendrimer layers than in the SAMs regardless of the surface density of functionalized biotin. Concurrently, the dendrimer layers led to much higher values of sticking probability, which is defined as the probability that the SA molecule adsorbs upon collision with a biotinylated surface, at a fixed SA coverage, and prolonged the significant levels around the maximum probability with increasing SA coverage. Plots of the saturation coverage of SA versus the SA concentration in solution showed that SA binding onto the biotinylated G1 and G3 layers fit to a Langmuir isotherm model. Taken together, faster binding of SA and highly ordered packing of the molecules seems to be achieved through typical properties of the dendrimer monolayers such as surface distribution of functionalized biotin, surface corrugation, and flexibility of highly branched larger dendrimers, which provides a guideline for the construction and analysis of an interfacial layer in biosensing applications.
Biotin↗
COMPAM :visualization of combining pairwise alignments for multiple genomes.
UNLABELLED: COMPAM is a tool for visualizing relationships among multiple whole genomes by combining all pairwise genome alignments. It displays shared conserved regions (blocks) and where these blocks occur (edges) as block relation graphs which can be explored interactively. An unannotated genome, e.g. can then be explored using information from well-annotated genomes, COG-based genome annotation and genes. COMPAM can run either as a stand-alone application or through an applet that is provided as service to PLATCOM, a toolset for whole genome comparative analysis, where a wide variety of genomes can be easily selected. Features provided by COMPAM include the ability to export genome relationship information into file formats that can be used by other existing tools. AVAILABILITY: http://bio.informatics.indiana.edu/projects/compam/
Algorithms↗
Inhibition assay of biomolecules based on fluorescence resonance energy transfer (FRET) between quantum dots and gold nanoparticles.
An inhibition assay method was developed based on the modulation in the FRET efficiency between quantum dots (QDs) and gold nanoparticles (AuNPs) in the presence of the molecules which inhibit the interactions between QD- and AuNP-conjugated biomolecules. For the functionalization, AuNPs were first stabilized by chemisorption of n-alkanethiols and then capped with the first generation polyamidoamine (G1 PAMAM) dendrimers. By employing a streptavidin-biotin couple as a model system, avidin was quantitatively analyzed as an inhibitor by sensing the change in photoluminescence (PL) quenching of SA-QDs by biotin-AuNPs. The detection limit for avidin was about 10 nM. It is anticipated that the PL quenching-based sensing system can be used for the quantitative analysis and high throughput screening of molecules which inhibit the specific biomolecular interactions.
Avidin↗
Mass spectrometric analysis of affinity-captured proteins on a dendrimer-based immunosensing surface: investigation of on-chip proteolytic digestion.
The monolayer of fourth-generation poly(amidoamine) dendrimers was adopted to construct the immunoaffinity surface of an antibody layer. The antibody layer as a bait on the dendrimer monolayer was found to result in high binding capacity of antigenic proteins and a reliable detection. The affinity-captured protein at the immunosensing surface was subjected to direct on-chip tryptic digestion, and the resulting proteolytic peptides were analyzed by using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The performance of the on-chip digestion procedure was investigated with respect to the ratio of trypsin to protein, digestion time, composition of a reaction buffer, and the amount of affinity-captured protein on a surface. Addition of a water-miscible organic solvent to a reaction buffer had no significant effect on the digestion efficiency under the optimized digestion conditions. The on-chip digestion method identified the affinity-captured bovine serum albumin (BSA), lysozyme, and ferritin at the level of around 100 fmol. Interestingly, the detected number of peptide hits through the on-chip digestion was almost similar regardless of the amount of captured protein ranging from low- to high-femtomole levels, whereas the efficiency of in-solution digestion decreased significantly as the amount of protein decreased to low-femtomole levels. The structural alignment of the peptide fragments from on-chip-digested BSA revealed that the limited exterior of the captured protein is subjected to attack by trypsin. The established detection procedures enabled the identification of BSA in the biological mixtures at the level of 0.1 ng/mL. The use of antibodies against the proteins involved in the metabolic pathway of L-threonine in Escherichia coli also led to discrimination of the respective target proteins from cell lysates.
Amino Acid Sequence↗