Robert Elghanian

Nanoparticle-based bio-barcode assay redefines “undetectable” PSA and biochemical recurrence after radical prostatectomy

We report the development of a previously undescribed gold nanoparticle bio-barcode assay probe for the detection of prostate specific antigen (PSA) at 330 fg/mL, automation of the assay, and the results of a clinical pilot study designed to assess the ability of the assay to detect PSA in the serum of 18 men who have undergone radical prostatectomy for prostate cancer. Due to a lack of sensitivity, available PSA immunoassays are often not capable of detecting PSA in the serum of men after radical prostatectomy. This new bio-barcode PSA assay is ≈300 times more sensitive than commercial immunoassays. Significantly, with the barcode assay, every patient in this cohort had a measurable serum PSA level after radical prostatectomy. Patients were separated into categories based on PSA levels as a function of time. One group of patients showed low levels of PSA with no significant increase with time and did not recur. Others showed, at some point postprostatectomy, rising PSA levels. The majority recurred. Therefore, this new ultrasensitive assay points to significant possible outcomes: (i) The ability to tell patients, who have undetectable PSA levels with conventional assays, but detectable and nonrising levels with the barcode assay, that their cancer will not recur. (ii) The ability to assign recurrence earlier because of the ability to measure increasing levels of PSA before conventional tools can make such assignments. (iii) The ability to use PSA levels that are not detectable with conventional assays to follow the response of patients to adjuvant or salvage therapies.

p24 antigen rapid test for diagnosis of acute pediatric HIV infection.

Currently, the majority of HIV-infected infants are found within limited-resource settings, where inadequate screening for HIV due to the lack of access to simple and affordable point-of-care tests impedes implementation of antiretroviral therapy. Here we report development of a low-cost dipstick p24 antigen assay using a visual readout format that can facilitate the diagnosis of HIV for infants in resource-poor conditions. A heat shock methodology was developed to optimize disruption of immune complexes present in the plasma of infected infants. The analytical sensitivity of the assay using recombinant p24 antigen is 50 pg/mL (2 pM) with whole virus detection as low as 42.5k RNA copies per milliliter plasma. In a blinded study comprising 51 archived infant samples from the Women and Infants Transmission Study, our assay demonstrated an overall sensitivity and specificity of 90% and 100%, respectively. In field evaluations of 389 fresh samples from South African infants, a sensitivity of 95% and specificity of 99% was achieved. The assay is simple to perform, requires minimal plasma volume (25 μL), and yields a result in less than 40 minutes making it ideal for implementation in resource-limited settings.

Use and evaluation of a [2+2] photoaddition in immobilization of oligonucleotides on a three-dimensional hydrogel matrix

Photochemical attachment of synthetic oligonucleotides on the three dimensional surface of a polyacrylamide based hydrogel was used in the specific detection of target oligonucleotides. Covalent attachment of the oligonucleotide to the hydrogel was mediated by the incorporation of a 2+2 photo-attachable functional group in both the hydrogel and the oligonucleotide probe. Expression and SNP assays were used to evaluate this platform.

CHEMICAL AND PHOTOCHEMICAL LIGATION OF OLIGONUCLEOTIDE BLOCKS

Abstract Several efficient means for joining oligonucleotides in dilute solution by non-natural internucleotide bridges are discussed. It is also shown that an oligonucleotide containing a -OP(O)(O−)S- link can hnction as an effective template in PCR amplification and that oligonucleotide probes containing stilbenedicarboxamide groups can serve in monitoring the presence of mismatched bases in an oligonucleotide target.

Membrane-based plasma collection device for point-of-care diagnosis of HIV

A major requirement for the development of point-of-care tests for the detection of disease analytes is the need to separate plasma from whole blood in an efficient and rapid manner. Furthermore, the separated plasma must be able to elute efficiently the analyte of interest and serve effectively as a physical matrix to deliver the equivalent of neat plasma for downstream diagnostic analysis. Additionally, many applications require the use of heat shock to liberate immunocomplexed antigen found in the collected plasma. A membrane-based filter method is reported for rapid and efficient collection of plasma from a whole blood sample that is compatible with heat shock. Using pediatric human immunodeficiency virus as an example, this device elutes 100% of the input p24 core antigen post-collection and enables heat shock of plasma samples identical to neat plasma treatment.

Chemistry of Oligonucleotide-Gold Nanoparticle Conjugates

Conjugates prepared by immobilizing thiol-terminated oligonucleotides onto gold nanoparticles form stable colloidal solutions in aqueous media. The oligonucleotides can serve as linkers to organize the gold particles reversibly into three dimensional assemblies, and the gold particles can function as colorimetric reporters for hybridization of the bound oligomers with target oligonucleotides in solution.

Dip pen nanolithography/spl trade/ and its potential for nanoelectronics

Dip pen nanolithography (DPN/spl trade/) is a patterning technique for nanoscale science and engineering based on scanning probe microscopy. Its main advantages are very high resolution, the unique capability to deposit many different materials directly onto a substrate and low cost of ownership. We present here new research and development efforts that demonstrate the potential of DPN as a tool to produce nanoelectronic devices and circuits. We show the direct deposition of electronic materials as well as the use of external accessories to accelerate the development phase of nanoelectronic components.