Sealed microwell assay
AssigneesCelldom, Inc., The Regents of the University of California
InventorsNatalia Tkachenko, Eugene Tkachenko, Alexander Groisman, Edgar Gutierrez
Life of this patent
Estimating the term…
Abstract
The present invention provides methods, systems and kits for performing microwell analysis in sealed small volumes. The microwells of the invention are sealed through the pairing of microwell and bead geometry wherein the diameter of the beads is greater than the diameter of the bottom of the microwells. Loading the beads into the microwells seals samples within an analytical space between the bottom of a nested bead and the bottom of the subject microwell.
Claims24
A kit for conducting an assay, comprising:
- a) a substrate having a plurality of microwells having a circular cross section;
- b) a plurality of beads comprising a ligand adapted to bind a target oligonucleotide;
- wherein said plurality of microwells and said plurality of beads are geometrically paired to permit said plurality of beads to (i) seal said plurality of microwells, and (ii) form an uninterrupted contact between a periphery of the plurality of beads and an inside wall of the plurality of microwells thereby providing a sealed partition beneath the plurality of beads when said plurality of beads are nested within said plurality of microwells;
- wherein each bead in said plurality of beads comprises at least one oligonucleotide comprising:
- (i) a first oligonucleotide portion forming said ligand;
- and (ii) a second oligonucleotide portion forming a bead label that is unique to each bead in said plurality of beads;
- wherein said first oligonucleotide portions are adapted to bind a plurality of different target oligonucleotides.
A method for single cell expression analysis, comprising:
- a) providing a substrate comprising a plurality of microwells having a circular cross section;
- b) loading cells into said plurality of microwells;
- c) contacting said plurality of microwells with a plurality of beads having a ligand that is adapted to bind an analyte, wherein (i) said plurality of microwells and said plurality of beads are geometrically paired to permit said plurality of beads to form an uninterrupted contact between a periphery of the plurality of beads and an inside wall of the plurality of microwells when said plurality of beads are contacted with said plurality of microwells, and (ii) contacting said plurality of microwells with said plurality of beads anchors said plurality of beads in said plurality of microwells thereby sealing said cells in said plurality of microwells beneath said uninterrupted contact;
- d) incubating said cells under conditions sufficient for said ligand to bind said analyte;
- and
- e) detecting binding of said ligand to said analyte thereby identifying the expression of said analyte by said cells.
Dependencies are inferred from the claim text.