# US 2014/0193315 A1 · Apparatus for loading molecules onto substrates

> Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt

- Number: US 2014/0193315 A1
- Document ID: US20140193315A1
- Status: Published application
- Kind: A1
- Application number: 14/149,402
- Filed: 2014-01-07
- Published: 2014-07-10
- Earliest claimed: 2011-03-23
- Assignees: Pacific Biosciences of California, Inc.
- Applicants: Pacific Biosciences of California, Inc.
- Inventors: Thang Pham; Jeremy Gray; Adrian Fehr; Aj Chmura; Yu-Chih Tsai; Arunashree Bhamidipati
- Web page: https://patentgrep.com/patent/US20140193315A1

## CPC

- C12Q 1/6874: involving nucleic acid arrays, e.g. sequencing by hybridisation
- B01L 9/523: {for multisample carriers, e.g. used for microtitration plates}
- C12Q 1/6869: Methods for sequencing
- G01N 33/54313: {the carrier being characterised by its particulate form}
- G01N 33/54353: {with ligand attached to the carrier via a chemical coupling agent (coatings G01N33/54393)}
- C40B 60/14: for creating libraries
- G01N 2333/922: Ribonucleases (RNAses); Deoxyribonucleases (DNAses)
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- B01L: CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)
- C40B: COMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES (in silico combinatorial libraries of nucleic acids, proteins or peptides G16B35/00; in silico combinatorial chemistry G16C20/60)

## Abstract

Disclosed are compositions, devices, and methods for loading molecules of interest onto a substrate by contacting beads having molecules of interest attached to them with the substrate, for example by providing a field that brings the beads into proximity or contact with the substrate and moves the beads with respect to the substrate. Such molecules of interest can be deposited onto substrates for single-molecule analysis.

## Claims

### Claim 1

An apparatus comprising;
a fixture for holding a substrate;
a substrate held within the fixture comprising an array of zero mode waveguides on its top surface, and comprising a reservoir for containing a solution in contact with the top surface of the substrate;
a solution, in contact with the substrate, comprising magnetic beads having molecules-of-interest attached thereto, the magnetic beads having diameters greater than the smallest cross-sectional dimensions of the zero mode waveguides; and
a device for generating a dynamic magnetic field disposed below, adjacent to, or above the substrate capable of generating a magnetic field that (i) pulls the magnetic beads to the top surface of the substrate, and (ii) moves the magnetic beads across the top surface of the substrate, whereby molecules-of-interest are deposited within the zero mode waveguides on the substrate.

### Claim 2 (depends on 1)

The apparatus of claim 1 wherein the molecules-of-interest are bound to the magnetic bead via hybridization between an oligonucleotide attached to the magnetic bead and a sequence on the template nucleic acid.

### Claim 3 (depends on 1)

The apparatus of claim 1 wherein the molecules-of-interest comprise polymerase-nucleic acid complexes.

### Claim 4 (depends on 1)

The apparatus of claim 1 wherein the magnetic bead is attached to a hook oligonucleotide comprising a retrieval sequence that is complimentary to an oligonucleotide attached to the magnetic bead and a capture sequence that is complementary to the template nucleic acid.

### Claim 5 (depends on 4)

The apparatus of claim 4 wherein the oligonucleotide attached to the magnetic bead comprises a poly(dA), poly(A), poly(dT) or poly(T) sequence.

### Claim 6 (depends on 4)

The apparatus of claim 4 wherein the relative binding strength of each of i) the magnetic bead to the hook oligonucleotide, ii) the hook oligonucleotide to the template nucleic acid, and iii) the polymerase-nucleic acid complex to the substrate are controlled such that when the polymerase-nucleic acid complex becomes bound to the substrate while applying the dynamic magnetic field, an attachment between the hook oligo and the template nucleic acid is broken.

### Claim 7 (depends on 1)

The apparatus of claim 1 wherein the dynamic magnetic field is produced using one or more moving permanent magnets.

### Claim 8 (depends on 1)

The apparatus of claim 1 wherein the dynamic field is produced using one or more electromagnets.

### Claim 9 (depends on 1)

The apparatus of claim 1 wherein the zero mode waveguides are cylindrical.

### Claim 10 (depends on 1)

The apparatus of claim 1 wherein the bases of the zero mode waveguides comprise a coupling agent.

### Claim 11 (depends on 10)

The apparatus of claim 10 wherein the coupling agent comprises biotin.

### Claim 12 (depends on 11)

The apparatus of claim 11 wherein the molecules-of-interest comprise polymerase-nucleic acid complexes, and the polymerase-nucleic acid complexes are attached to a biotin binding protein which binds to the coupling agent.

### Claim 13 (depends on 12)

The apparatus of claim 12 wherein the biotin binding protein comprise streptavidin, neutravidin, or avidin.