Scientific Publications

Nanoparticle-probe-based, Automated Assay for Ultra-sensitive and Multiplexed Diagnostic Applications

Oak Ridge Conference (AACC)
April 14, 2011

Our aims were to develop the ultra-sensitivity of nanoparticle probe based automated assay for multiplexed protein and DNA detection for high throughput diagnostic applications.

The nanoparticle probe based assay format is broadly applicable across various protein biomarkers and uses a multi-step deterministic robotic process, which relies on non-isotropically oriented antibodies on functionalized glass as multiplexed microarrays to capture protein or DNA markers from biological samples. Functionalized, 130 angstrom diameter gold-probes also bind to the markers through a molecular-scale complex containing antibodies or DNA hybridization. The marker-bound molecular complex is then quantified through silver enhancement of the functionalized gold.

We have demonstrated very robust assays with troponin I detection (LOD < 200fg/ml), rheumatoid arthritis (RA) tests, allergen detection, PSA detection (LOD < 100fg/mL) and other cancer markers including NQO1, uteroglobin, CLC-5, uroplakin, RSOR, p53, CA125, AFP, HE4, CA15-3, CA19-9, beta-hCG, and CEA.

The clinical applications have been demonstrated by testing over 1500 serum samples for cTnI assay, 1500 serum samples for RA tests, 250 allergen samples, 288 bladder cancer serum samples, 144 kidney cancer samples, and over 500 PSA serum samples. The ultra-sensitive gold nano-particle probe-based assay could provide a powerful tool for evaluating novel tumor markers, validation and clinical applications, including early diagnosis.

Ultrasensitive and multiplexed cancer marker detection via nanoparticle probes-based microarray assay

American Society of Clinical Oncology (ASCO)
October 19, 2010

Background: Aiming to improve the sensitivity of tumor marker assays, we have developed a microarray based nano-probe system and assays.

Methods: Our tumor marker assay format is broadly applicable across various protein biomarkers and uses a multistep deterministic robotic process, which relies on non-isotropically oriented antibodies on functionalized glass as multiplexed microarrays to capture tumor markers from serum. Functionalized, 130 angstrom diameter gold-probes also bind to the tumor markers through a molecular scale complex containing antibodies. The tumor marker-bound molecular complex is then quantified through silver enhancement of the functionalized gold.

Results: Our test array contains captures for putative tumor biomarkers for bladder, ovarian, prostate and renal cancers. We have demonstrated a very robust assay with sub-picomolar sensitivity, an overall CV less than 20%, and high clinical sensitivity and specificity. For example, the LOD of PSA is less than 100 femtograms per mL serum. The assay also showed very low background, broad dynamic range and over 3 logs of linear dose response.

Conclusions: The ultra-sensitive gold nano-particle probe-based tumor marker assay could provide a powerful tool for evaluating novel tumor markers, validation and clinical applications, including early diagnosis.

Nanoparticle-based ultra-sensitive assay for discovery and evaluation of cancer markers

J Clin Oncol 29: 2011 (suppl; abstr e21169)
May 18, 2011

Background: Our aim was to develop ultra-sensitive nanoparticle based assay for discovery of new cancer markers and evaluation of diagnostic applications.

Methods: The nanoparticle probe based assay format uses a multi-step deterministic robotic process, which relies on non-isotropically oriented antibodies on functionalized glass as multiplexed microarrays to capture protein or DNA markers from biological samples. Functionalized, 130 angstrom diameter gold-probes also bind to the markers through a molecular-scale complex containing antibodies or DNA hybridization. The marker-bound molecular complex is then quantified through silver enhancement of the functionalized gold.

Results: We have demonstrated the feasibility for the detection of cancer markers including NQO1, uteroglobin, CLC-5, uroplakin, RSOR, p53, CA125, AFP, HE4, CA15-3, CA19-9, beta-hCG, CEA, and PSA. The high sensitivity (e.g., PSA LOD <0.1pg/mL, uteroglobin LOD <10pg/mL) and broad dynamic range were demonstrated by target titrations in both buffer and serum samples. The clinical application has been explored by testing 288 bladder cancer serum samples, 144 kidney cancer samples, and over 500 PSA serum samples.

Conclusions: The ultra-sensitive gold nano-particle probe-based tumor marker assay could provide a powerful tool for evaluating novel tumor markers, validation and clinical applications, including early diagnosis.

Tumor Marker Detection by Nanofishing Test using Ultra Sensitive Nanoassay

Abbott Symposium: Advances in Tumor Immunology
December 2, 2011
Abstract (to accompany poster):

Aiming to improve the performance of tumor marker assays, earlier we have developed a microarray based nano-probe system. Our tumor assay format is broadly applicable across various protein biomarkers and uses a multi-step deterministic robotic process, which relies on non-isotropically oriented antibodies on functionalized glass as multiplexed microarrays to capture tumor markers from serum or plasma. Functionalized gold probes, which are in the order of a few hundred angstroms diameter, also bind to the tumor markers through a molecular-scale complex containing antibody. The tumor marker-bound molecular complex is then quantified through silver enhancement of the functionalized gold.

Based on the ultrasensitive nano-assay system, we have developed a novel assay format for detection of tumor markers for prostate cancer which utilizes the autoantibodies present in patient serum or plasma. This format is coined as ‘nanofishing’ test. This new assay format was characterized and compared to the conventional mode of tumor marker detection. Based on the ultra-sensitive gold nano-particle probe to detect tumor markers, such nanofishing assays could provide a powerful tool for evaluating novel biomarkers, validation and clinical applications, including early diagnosis and prognosis.

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