Leland Stanford Junior University

University of California

Magarray, Inc.

Nader Pourmand

Ronald W Davis

Miloslav Karhanek

Paolo Actis

Shan X Wang

Boaz Vilozny

R Adam Seger

Senkei Umehara

Chris David Webb

Robert L White

Heng Yu

Arjang Hassibi

Sebastian J Osterfeld

Bakthan Singaram

Baback Gharizadeh

Lisa Diamond

Jochen Kumm

Michelle M Maalouf

Muhammad Akram Tariq

Provided are methods for simultaneously detecting analytes. The methods include exposing a tip of a nanosensor to a biological sample, measuring ionic current flow through the nanosensor tip to detect a first analyte and a second analyte in the biological sample, and distinguishing the first analyte from the second analyte based on a first diffusion limited current peak resulting from binding of a first specific binding member to the first analyte being distinguishable from a second diffusion limited current peak resulting from binding of a second specific binding member to the second analyte. Sensing apparatuses that find use, e.g., in practicing the methods of the present disclosure, are also provided.

Methods for simultaneous detection of analytes and apparatuses for practicing same

Methods and apparatus for direct detection of chemical reactions are provided. Electric charge perturbations of the local environment during enzyme-catalyzed reactions are sensed by an electrode system with an immobilized target molecule. The charge perturbation caused by the polymerase reaction can uniquely identify a DNA sequence. The polymerization process generates local perturbations of charge in the solution near the electrode surface and induces a charge in a polarazible gold electrode. This event is detected as a transient current by a voltage clamp amplifier. Detection of single nucleotides in a sequence can be determined by dispensing individual dNTPs to the electrode solution and detecting the charge perturbations. Alternatively, multiple bases can be determined at the same time using a mix of all dNTPs with subsequent analysis of the resulting signal. This technique may be adapted to other reaction determinations, such as enzymatic reactions, other electrode configurations, and other amplifying circuits.

Charge perturbation detection system for DNA and other molecules

Disclosed are methods and devices for biomolecular detection, comprising a nanopipette, exemplified as a hollow inert, non-biological structure with a conical tip opening of nanoscale dimensions, suitable for holding an electrolyte solution which may contain an analyte such as a protein biomolecule to be detected as it is passed through the tip opening. Biomolecules are detected by specific reaction with peptide ligands chemically immobilized in the vicinity of the tip. Analytes which bind to the ligands cause a detectible change in ionic current. A sensitive detection circuit, using a feedback amplifier circuit, and alternating voltages is further disclosed. Detection of IL-10 at a concentration of 4 ng/ml is also disclosed, as is detection of VEGF.

Functionalized nanopipette biosensor

Disclosed are methods and devices for detection of ion migration and binding, utilizing a nanopipette adapted for use in an electrochemical sensing circuit. The nanopipette may be functionalized on its interior bore with metal chelators for binding and sensing metal ions or other specific binding molecules such as boronic acid for binding and sensing glucose. Such a functionalized nanopipette is comprised in an electrical sensor that detects when the nanopipette selectively and reversibly binds ions or small molecules. Also disclosed is a nanoreactor, comprising a nanopipette, for controlling precipitation in aqueous solutions by voltage-directed ion migration, wherein ions may be directed out of the interior bore by a repulsing charge in the bore.

Nanopore device for reversible ion and molecule sensing or migration

Methods for analyte detection with magnetic sensors are provided. Aspects of the methods include producing a magnetic sensor device having a magnetically labeled analyte from a sample, such as a serum sample, bound to a surface of a magnetic sensor thereof; and obtaining a signal, e.g., a real-time signal, from the magnetic sensor to determine whether the analyte is present in the sample. Also provided are devices, systems and kits that find use in practicing the methods of the invention. The methods, devices, systems and kits of the invention find use in a variety of different applications, including detection of biomarkers, such as disease markers.

Analyte detection with magnetic sensors

Described herein are devices and methods for extracting cellular material from living cells and then depositing them into to a receptacle in a nanoliter scale. Using a nanopipette integrated into a scanning ion conductance microscope (SICM), extraction of mitochondrial DNA from human BJ fibroblasts and Green Fluorescent Protein (GFP) transcripts from HeLa/GFP cells was achieved with minimal disruption to the cellular milieu and without chemical treatment prior to obtaining the isolated sample. Success of the extraction was confirmed by fluorescence microscopy and PCR analysis of the extracted material. The method and apparatus may be applied to many different cell types and intracellular targets, allowing not only single cell analysis, but single subcellular compartment analysis of materials extracted in their native state.

Nanopipette device and method for subcellular analysis

Disclosed herein are methods and systems for controlled ejection of desired material onto surfaces including in single cells using nanopipettes, as well as ejection onto and into cells. Some embodiments are directed to a method and system comprising nanopipettes combined with an xyz controller for depositing a user defined pattern on an arbitrary substrate for the purpose of controlled cell adhesion and growth. Alternate embodiments are directed to a method and system comprising nanopipettes combined with an xyz controller and electronic control of a voltage differential in a bore of the nanopipette electroosmotically injecting material into a cell in a high-throughput manner and with minimal damage to the cell. Yet other embodiments are directed to method and system comprising functionalized nanopipettes combined with scanning ion conductance microscopy for studying molecular interactions and detection of biomolecules inside a single living cell.

Nanopipette apparatus for manipulating cells

Disclosed are methods and devices for biomolecular detection, comprising a nanopipette, exemplified as a hollow inert, non-biological structure with a conical tip opening of nanoscale dimensions, suitable for holding an electrolyte solution which may contain an analyte such as a protein biomolecule to be detected as it is passed through the tip opening. Biomolecules are detected by specific reaction with peptide ligands chemically immobilized in the vicinity of the tip. Analytes which bind to the ligands cause a detectable change in ionic current. A sensitive detection circuit, using a feedback amplifier circuit, and alternating voltages is further disclosed. Detection of IL-10 at a concentration of 4 ng/ml is also disclosed, as is detection of VEGF.

Growing community of inventors

San Mateo, CA, United States of America

Nader Pourmand