Tagged ligands for enrichment of rare analytes from a mixed sample

• US 8,008,032 B2
• Filed: 02/25/2008
• Issued: 08/30/2011
• Est. Priority Date: 02/25/2008
• Status: Expired due to Fees

• Current Assignee: ZEON Corporation
• Original Assignee: CELLective Dx Corporation
• Inventors: Barnes, Helen, Forsyth, Allyn
• Primary Examiner(s): GABEL, GAILENE
• Application Number:
  US12/037,077
• Publication Number:
  US 20090215088A1
• Time in Patent Office: 1,282 Days
• Field of Search: 435/2, 435/7.2, 435/7.23, 435/7.5, 435/40.51, 435/286.6, 435/287.1, 435/971, 435/973, 435/286.5, 435/287.2, 436/501, 436/518, 436/56, 436/64, 436/167, 436/175, 436/177, 436/813, 436/524, 436/528, 436/532, 436/10, 422/101, 422/417, 422/502, 422/504
• US Class Current: 435/7.23
• CPC Class Codes: B01L 2200/0647 Handling flowable solids, e.g. microscopic beads, cells, particles
• CPC Class Codes: B01L 2300/0861 Configuration of multiple channels and/or chambers in a single devices
• CPC Class Codes: B01L 2400/086 using baffles or other fixed flow obstructions
• CPC Class Codes: B01L 3/502761 specially adapted for handling suspended solids or molecules independently from the bulk fluid flow, e.g. for trapping or sorting beads, for physically stretching molecules investigating characteristics of particles G01N15/00
• CPC Class Codes: G01N 1/405 by adsorption or absorption
• CPC Class Codes: G01N 2333/705 Assays involving receptors, cell surface antigens or cell surface determinants
• CPC Class Codes: G01N 33/54326 Magnetic particles
• CPC Class Codes: G01N 33/574 for cancer
• CPC Class Codes: G01N 33/57492 involving compounds localized on the membrane of tumor or cancer cells
• CPC Class Codes: G01N 33/585 with a particulate label, e.g. coloured latex
• CPC Class Codes: Y10S 435/971 Capture of complex after antigen-antibody reaction
• CPC Class Codes: Y10S 435/973 Simultaneous determination of more than one analyte
• CPC Class Codes: Y10T 436/101666 Particle count or volume standard or control [e.g., platelet count standards, etc.]
• CPC Class Codes: Y10T 436/13 Tracers or tags
• CPC Class Codes: Y10T 436/25125 Digestion or removing interfering materials
• CPC Class Codes: Y10T 436/25375 Liberation or purification of sample or separation of material from a sample [e.g., filtering, centrifuging, etc.]

Abstract

Method of enriching specific cells from cellular samples are disclosed, comprising contacting in solution a cellular sample with affinity-tagged ligands (ATLs) each comprising a first ligand linked to an affinity tag, wherein the ligand selectively binds a cellular marker of the rare cells and the affinity tag can be selectively captured by a capture moiety, wherein the affinity tags do not comprise a magnetic particle; and flowing the sample through a microfluidic device comprising the capture moiety to selectively retain ATL-bound cells. Methods for enriching circulating tumor cells, and devices for enriching specific cells from cellular samples are also disclosed.

Claims

1. A method for enriching rare cells from a cellular sample comprising:

(a) contacting in solution said cellular sample with affinity-tagged ligands (ATLs) each ATL comprising;

a first ligand linked to an affinity tag, wherein said ligand selectively binds a cellular marker of said rare cells, said affinity tag can be selectively captured by a capture moiety, and said affinity tag does not comprise a magnetic particle; and

(b) flowing said cellular sample containing said ATLs through a microfluidic device comprising a substrate coated with said capture moiety to thereby selectively retain ATL-bound cells by binding said affinity tag to said capture moiety.

2. The method of claim 1, wherein said rare cells are circulating tumor cells or epithelial cells.

3. The method of claim 1, wherein said ligand is folic acid, epidermal growth factor (EGF), Dolichos Biflorus Agglutinin (DBA), EGF peptide, epi peptide, transforming growth factor-alpha (TGF-alpha), Urokinase Plasminogen Activator (UPA), FasL, MUC1/sec, catenin, ICAM-1, or plasminogen.

4. The method of claim 1, wherein said ligand is an antibody.

5. The method of claim 4, wherein said antibody is anti-EpCam, anti-ErbB2, or anti-CK19 antibody.

6. The method of claim 1, wherein said ligand is not an antibody.

7. The method of claim 1, wherein said affinity tag is biotin, desthiobiotin, histidine, polyhistidine, myc, hemagglutinin (HA), FLAG, fluorescence tag, tandem affinity purification (TAP) tags, or glutathione S transferase (GST) or derivatives thereof.

8. The method of claim 1, wherein said ATLs comprises:

(a) a first ATL comprising a first ligand that selectively binds a first cellular marker of the rare cells., wherein said first ligand is linked to a first affinity tag, and (b) a second ATL comprising a second ligand that selectively binds a second cellular marker of the rare cells, wherein said second ligand is linked to a second affinity tag, wherein the first ligand and the second ligand are different.

9. The method of claim 1, wherein said ATLs comprises at least 3, 4, 5, 6, 7, or 8 different ATLs, each comprising a ligands, which selectively binds a different cellular marker of the rare cells.