US 6,919,183 B2 · Isolating cells expressing secreted proteins

Abstract

A method for identifying and isolating cells which produce secreted proteins. This method is based upon a specific characteristic or the expression level of the secreted protein by transiently capturing the secreted protein on the surface of an individual cell, allowing selection of rare cell clones from a heterogeneous population. Also provided is the use of this method to generate cells which produce a desired level of secreted protein or secreted protein of a particular characteristic(s), and organisms which possess such cells. In particular, the method allows rapid isolation of high expression recombinant antibody-producing cell lines, or may be applied directly to rapid isolation of specific hybridomas, or to the isolation of antibody-producing transgenic animals. This method is applicable for any cell which secretes protein.

Claims47

Dependent claims

A method of detecting and/or isolating a eukaryotic cell that produces a secreted protein of interest (POI), comprising:

(a) providing a cell comprising a nucleic acid that encodes a secreted POI and a nucleic acid that encodes a cell surface capture molecule capable of binding the POI, wherein the nucleic acid encoding the secreted POI or the nucleic acid encoding the cell surface capture molecule, or both, are transfected into the cell;
(b) culturing the cell under conditions in which the POI and cell surface capture molecule are expressed, and a POI-capture molecule complex is formed intracellularly and displayed on the cell surface;
(c) contacting the cell with a detection molecule, which binds to the POI displayed by the cell;
and
(d) detecting and/or isolating the cell due to it being bound to the detection molecule.

2 more elements

Dependencies are inferred from the claim text.

Isolating cells expressing secreted proteins

Life of this patent

Isolating cells expressing secreted proteins

Life of this patent

Abstract

Claims47