Analysis Of Rare Cell-Enriched Samples
The present invention relates to methods for detecting, enriching, and analyzing rare cells that are present in the blood, e.g., epithelial cells. The invention further features methods of analyzing rare cell(s) to determine the presence of an abnormality, disease or condition in a subject by analyzing a cellular sample from the subject.
1 . A method for detecting cancer in a subject comprising:
enriching a sample from said subject for rare cells by flowing said sample through an array of obstacles coated with antibodies that specifically bind to one or more cell populations in said sample to obtain a rare cell-enriched sample, wherein said rare cells in said sample are in a concentration of less than 1 in 100,000 cells prior to said enrichment, and
detecting the presence or absence of a rare cell nucleic acid in said rare cell-enriched sample, wherein the presence of said rare cell nucleic acid in said rare cell-enriched sample indicates the presence of said cancer in said subject.
2 . The method of claim 1 , wherein said array of obstacles comprises about 5,000 to about 100,000 obstacles.
3 . The method of claim 1 , wherein said rare cell-enriched sample is not eluted from said array prior to said detecting.
4 .- 6 . (canceled)
7 . The method of claim 1 , wherein said sample is a blood sample.
8 . (canceled)
9 . The method of claim 1 , wherein said rare cells comprise circulating tumor cells, epithelial cells, endothelial cells, or progenitor stem cells.
10 . (canceled)
11 . The method of claim 9 , wherein said detecting comprises detecting the expression of a gene identified in FIG. 5 .
12 . The method of claim 11 , further comprising detecting the presence or absence of a mutation in said gene.
13 . The method of claim 11 , wherein a level of expression of said gene is determined.
14 . The method of claim 11 , wherein said gene is EGFR, EGF, EpCAM, MUC-1, HER-2, or Claudin-7.
15 .- 18 . (canceled)
19 . The method of claim 1 , wherein said antibodies comprise an anti-EpCAM antibody, an anti-E-cadherin antibody, or an anti-Muc-1 antibody.
20 .- 21 . (canceled)
22 . The method of claim 1 , wherein said antibodies comprise an anti-CD71 antibody, anti-CD235a antibody, anti-CD36 antibody, an anti-selectin antibody, an anti-CD45 antibody, or an anti-GPA antibody.
23 . (canceled)
24 . The method of claim 1 , further comprising flowing said sample or said enriched sample through an array of obstacles that selectively directs cells larger than a predetermined size in to a first outlet and cells equal to or smaller than said predetermined size to a second outlet.
25 . (canceled)
26 . A method for detecting cancer in a subject comprising:
(i) enriching a sample from said subject for rare cells by flowing said sample through an array of obstacles that selectively directs cells smaller than a predetermined size to a first outlet and cells equal to or larger than said predetermined size to a second outlet to obtain a first rare cell-enriched sample, wherein said rare cells in said sample are in a concentration of less than 1 in 100,000 cells;
(ii) flowing said first rare cell-enriched sample through an array of obstacles coated with antibodies that specifically bind to one or more cell populations in said sample to obtain a second rare cell-enriched sample; and
(iii) detecting the presence or absence of a rare cell nucleic acid in said second rare-cell enriched sample, wherein the presence of said rare cell nucleic acid in said second rare-cell enriched sample indicates the presence of said cancer in said subject.
27 . The method of claim 26 , wherein said predetermined size is about the size of a red blood cell, a white blood cell, a circulating tumor cell, an epithelial cell, an endothelial cell, or a progenitor stem cell.
28 . The method of claim 26 , wherein said predetermined size is about 1 to about 10 μm.
29 . The method of claim 26 , wherein said rare cell nucleic acid is from an epithelial cell.
30 . The method of claim 26 , wherein said rare cell nucleic acid is EGFR, EGF, EpCAM, MUC-1, HER-2, or Claudin-7.
31 .- 32 . (canceled)
33 . The method of claim 26 , wherein said cancer is breast cancer, lung cancer, or prostate cancer.
34 .- 42 . (canceled)
43 . A kit for detecting cancer cells in a subject, comprising a device comprising an array of obstacles coated with antibodies that specifically bind to one or more cell populations and a set of reagents for detecting the expression of a gene identified in FIG. 5 .
44 . The kit of claim 43 , wherein said gene is EGFR, EGF, EpCAM, MUC-1, HER-2, or Claudin-7.
45 .- 47 . (canceled)