Efficient enrichment and detection of disseminated tumor cells

Abstract

Tumor cells, particularly carcinoma cells, are separated from peripheral blood by magnetic sorting. The tumor cells are magnetically labeled with antibodies directed to tissue specific antigens, preferably cytoplasmic proteins. Labeling for cytoplasmic antigens is accomplished first permeabilizing, then fixing the cells. The cells are separated on a magnetic matrix. The number of tumor cells in the enriched fraction is used to calculate the number of tumor cells present in a patient hematopoietic sample.

What is claimed is:  
     
         1 . A method for separation of target cells from a whole blood cell sample present as a fluid suspension, wherein said target cells express one or more antigens in a tissue specific manner, wherein said antigens are absent on the majority of blood cells, the method comprising the steps, in order, of: 
 adding to said whole blood cell sample antibodies specific for said antigens, wherein said antibodies are coupled to a magnetically responsive reagent, under conditions sufficient to specifically bind said coupled antibodies to said antigens;    passing said sample through a ferromagnetic matrix in the presence of a magnetic field to magnetically immobilize cells in the sample having coupled antibodies bound thereto;    washing said matrix of unbound cells; and    removing the magnetic field and eluting bound cells from said matrix directly onto a solid support to provide an enriched cell sample comprising target cells, wherein said eluting is by centrifugation or vacuum filtration.    
     
     
         2 . The method of    claim 1   , wherein said solid support is a polycarbonate filter or a slide.  
     
     
         3 . The method of    claim 1   , wherein said coupled antibodies are specific for one or more antigens selected from the group consisting of epithelial membrane antigen, human embryonic antigen, human milk fat globules, MBr1, MBr8, Ber-EP4, 17-1A, C26, T16, desmin, muscle-specific actin, placental alkaline phosphatase, beta-human chorionic gonadotropin, alpha-fetoprotein, prostate specific antigen, carcinoembryonic antigen, HMB-45, chromagranin-A, and synaptophysin.  
     
     
         4 . A method for separation of non-hematopoietic target cells from a whole blood cell sample present as a fluid suspension, wherein said target cells comprise more than one cell per 10 7  blood cells, the method comprising the steps, in order, of: 
 adding to said whole blood cell sample antibodies specific for one or more antigens expressed by said target cells and absent on blood cells, wherein said antibodies are coupled to a magnetically responsive reagent, under conditions sufficient to specifically bind said coupled antibodies to said antigens;    passing said sample through a ferromagnetic matrix in the presence of a magnetic field to magnetically immobilize target cells in the sample having coupled antibodies bound thereto;    washing said matrix of unbound cells; and    removing the magnetic field and eluting bound cells from said matrix directly onto a solid support to provide an enriched cell sample comprising non-hematopoietic target cells, wherein said eluting is by centrifugation or vacuum filtration.    
     
     
         5 . The method of    claim 4   , wherein said solid support is a polycarbonate filter or a slide.  
     
     
         6 . The method of    claim 4   , wherein said coupled antibodies are specific for one or more antigens selected from the group consisting of epithelial membrane antigen, human embryonic antigen, human milk fat globules, MBr1, MBr8, Ber-EP4, 17-1A, C26, T16, desmin, muscle-specific actin, placental alkaline phosphatase, beta-human chorionic gonadotropin, alpha-fetoprotein, prostate specific antigen, carcinoembryonic antigen, HMB-45, chromagranin-A, and synaptophysin.