Composite blastocysts (CBs) from aggregates of dissociated cells of non-viable pre-embryos

• US 6,759,244 B2
• Filed: 11/08/2001
• Issued: 07/06/2004
• Est. Priority Date: 11/08/2001
• Status: Expired due to Fees

• Current Assignee: Art Institute of New York and New Jersey Incorporated
• Original Assignee: Art Institute of New York and New Jersey Incorporated
• Inventors: Willadsen, Steen Malte, Alikani, Mina
• Primary Examiner(s): Paras, Peter
• Assistant Examiner(s): BERTOGLIO, VALARIE E
• Application Number:
  US10/036,581
• Publication Number:
  US 20030087431A1
• Time in Patent Office: 971 Days
• Field of Search: 435/373, 435/366, 435/363, 424/582
• US Class Current: 435/373
• CPC Class Codes: C12N 15/873 Techniques for producing new embryos, e.g. nuclear transfer, manipulation of totipotent cells or production of chimeric embryos
• CPC Class Codes: C12N 2503/04 Screening or testing on artificial tissues
• CPC Class Codes: C12N 5/0604 Whole embryos; Culture medium therefor
• CPC Class Codes: C12N 5/0606 Pluripotent embryonic cells, e.g. embryonic stem cells [ES] embryonic germ cells C12N5/0611, induced pluripotent stem cells C12N5/0696

Abstract

A preparation and a method of making composite blastocysts (CBs) from aggregates of dissociated cells of non-viable pre-embryos are disclosed. The CB is characterized morphologically by having two distinct tissue types, the inner cell mass (ICM) and the trophectoderm (TE), and a blastocoelic cavity (BC). The method of making CBs is an aggregation process (AP) comprising inter alia the following steps: 1) dissociation of discarded pre-embryos; 2) isolation of single nucleated cells from dissociated discarded pre-embryos; 3) microsurgical encapsulation of several cells within a host zona pellucida or artificial aggregation with or without a non-zona vessel; and 5) primary culture of the cell aggregates for multiplication and differentiation of cells. One particularly advantageous embodiment is that the starting material is non-viable pre-embryos. Another advantageous embodiment is that the AP allows individual cells from non-viable pre-embryos to further multiply, and become integrated into CBs. The novel CBs and the novel aggregation process disclosed.

Claims

1. A method of producing composite blastocysts comprising:

a) dissociation of non-viable mammalian pre-blastocyst embryos into non-nucleated and individual nucleated cells or groups of cells;

b) isolation of individual mononucleated cells or groups of mononucleated cells from disaggregated non-viable pre-blastocyst embryos;

c) aggregation of isolated mononucleated cells or groups of mononucleated cells from non-viable pre-blastocyst embryos in a host zona pellucida; and

d) culturing of the zona-encapsulated cell aggregates to allow multiplication of cells to form a composite blastocyst.