Graphite-Coated Magnetic Nanoparticle Microarray for Few-Cells Enrichment and Detection

Graphite-coated, highly magnetic FeCo core–shell nanoparticles were synthesized by a chemical vapor deposition method and solubilized in aqueous solution through a unique polymer mixture modification, which significantly improved the biocompatibility and stability of the magnetic nanoparticles (MNPs). Such functionalized MNPs were proven to be very stable in different conditions which would be significant for biological applications. Cell staining, manipulation, enrichment, and detection were developed with these MNPs. Under external magnetic manipulation, the MNP-stained cells exhibited directed motions. Moreover, MNPs were printed on substrates to modulate the magnetic field distribution on the surface. Capture and detection of sparse populations of cancer cells spiked into whole blood has been explored in a microarray fashion. Cancer cells from hundreds down to only two were able to be simply and efficiently detected from 1 mL of whole blood on the MNP microarray chips. Interestingly, the cells captured through the MNP microarray still showed viability and adhered to the MNP spots after incubation, which could be utilized for cancer cell detection, localized growth, and proliferation

Peptide-antigen microarrays on plasmonic gold substrate for profiling antibodies in serum samples of SLE patients and healthy individuals.

<p>A) Box plot of serum IgG antibody reactivity against several peptides and a whole antigen for 20 SLE patients and 20 healthy controls. Acetylated histone H2B peptides were found to be able to differentiate SLE patients and healthy controls (top box plots) together with whole H2B protein (bottom left box plot). While the H3 peptide with the 18th lysine methylated were found not capable of telling SLE patient from healthy control (bottom right box plot). B–D) Heatmaps displaying antibody reactivity to (B) histone peptides only, (C) whole antigens only, and (D) a combination of histone peptides and whole antigens that are identified capable of differentiating SLE patients and healthy controls with false discovery rate (q value) <0.001. The dashed lines are drawn to highlight the separation of SLE and healthy groups identified by using the average linkage Euclidean distance hierarchical clustering method. Color intensity of each grid in the heatmap reflected mean fluorescence intensity of corresponding peptide or antigen spot on the microarray for each SLE patient or healthy individual. In (B) and (C), several SLE patients (labeled in blue and purple color) are misplaced in the healthy group. These patients are grouped in the SLE side in (D) that profiles antibodies against both peptides and whole antigens. However, one healthy individual is mis-placed in the SLE group by this approach, reducing the specificity of this analysis.</p

An integrated peptide-antigen microarray for profiling serum antibodies of 20 patients with systemic lupus erythematosus (SLE).

<p>A) Peptide-antigen microarray layout including modified and unmodified histone H2A, H2B, H3, H4 peptides as well as whole antigens. Ac: acetylated; aa: amino acid; Me1: methylated; Me2: dimethylated; Me3: trimethylated; Ph: phosphorylated; K: Lysine; S: Serine. Number indicates amino acid position from the N-terminus of its corresponding histone proteins. Peptide sequences are listed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0071043#pone.0071043.s008" target="_blank">Table S1</a>. B) Microarray imaging results of a SLE patient serum probed on a plasmonic gold substrate (left) vs. on a commercial streptavidin-glass substrate (right) shown in the same IRDye800 fluorescence intensity scale. C) Dynamic range of fluorescence signal reading among 20 SLE patients measured on each peptide and antigen spots on avidin/gold slide (upper panel) and commercial streptavidin/glass slide (lower panel). Each dot represents a fluorescence intensity measured on a peptide or antigen spot shown along the x-axis in the serum of each of the 20 SLE patients. A dot is drawn only when the fluorescence intensity is above the background by 2 standard deviation of the background signal.</p

Plasmonic gold films for peptide microarrays.

<p>A) Scanning electron microscopy (SEM) of an as-made plasmonic gold film containing nano-islands formed on a glass substrate. Inset: an X-ray photoelectron spectrum (XPS) of the film. B) SEM of a plasmonic gold film coated with a layer of avidin proteins (avidin coating caused blurred features of the underlying gold nano-islands). Inset, an XPS spectrum taken on the avidin coated gold substrate showing the presence of nitrogen (in the avidin) over the film. C) A schematic drawing of a multiplexed peptide array (two peptides are shown, red and blue; peptide spots are surrounded by PEG-star coating) on gold for antibody screening based on a 3-layer assay with a NIR fluorophore reporter. D) Intensity of IRDye800 fluorescence (emission ∼ 800 nm) measured on a layer of IRDye800 labeled avidin coated on glass vs. on a plamonic gold film. A ∼100 fold fluorescence enhancement was observed on gold.</p