In-Vitro fertilization systems with spermatozoa separated into X-chromosome and Y-chromosome bearing populations

• US 20060281176A1
• Filed: 08/21/2006
• Published: 12/14/2006
• Est. Priority Date: 11/29/2000
• Status: Active Grant

• Current Assignee: XY, LLC (Inguran LLC)
• Original Assignee: Colorado State University (Colorado State University System), XY Incorporated (Inguran LLC)
• Inventors: Seidel, George E., Lu, Kehuan, Suh, Tae Kwang
• Granted Patent: US 8,137,967 B2
• Time in Patent Office: Days
• US Class Current: 435/325
• CPC Class Codes: A01N 1/02 Preservation of living parts
• CPC Class Codes: A01N 1/0263 Non-refrigerated containers specially adapted for transporting or storing living parts whilst preserving, e.g. cool boxes, blood bags or "straws" for cryopreservation
• CPC Class Codes: A01N 1/0273 Transport containers A01N1/0268 takes precedence
• CPC Class Codes: A61D 19/00 Instruments or methods for reproduction or fertilisation
• CPC Class Codes: C12N 5/0604 Whole embryos; Culture medium therefor

Abstract

An IVF system for successfully utilizing spermatozoa separated into X-chromosome bearing and into Y-chromosome bearing population for insemination. The IVF system includes fertilization medium that can shorten the time from insemination to cleavage and a portable incubator for the transportation of maturing oocytes and inseminated oocytes comprising a straw (19) and an incubation element (20) that can be sealed with a cap (22).

Claims

1. A method of fertilizing oocytes, comprising the steps of:

a. collecting sperm cells from a non-human male mammal;

b. transferring said sperm cells to a straw containing a plurality of oocytes in fertilization medium, wherein said oocytes are from a female mammal of the same species as said male mammal;

c. sealing said straw containing said sperm cells and said oocytes;

d. transferring said straw containing said sperm cells and said oocytes to the inside of an incubation element;

e. establishing incubation conditions within said incubation element;

f. sealing said incubation element; and

g. fertilizing at least one oocyte in said straw.

2. A method of transporting oocytes as described in claim 1, wherein said female mammal is selected from the group consisting of primates, bovids, ovids, equids, swine, and dolphins.

3. A method of transporting oocytes as described in claim 1, wherein said fertilization medium comprises modified Tyrode'"'"'s medium supplemented with 0.6 percent bovine serum albumin, 20 μ

g heparin per milliliter of Tyrode'"'"'s medium, and a concentration of 5 milli-molar caffeine.

4. A method of transporting oocytes as described in claim 1, wherein between about 10 and about 15 of said oocytes are contained within about 50 micro-liters of said fertilization medium.

5. A method of transporting oocytes as described in claim 1, wherein said straw has heat sealable aperture elements.

6. A method of transporting oocytes as described in claim 5, wherein said straw has a interior volume of about 0.25 milliliters.

7. A method of transporting oocytes as described in claim 1, wherein said incubation element has sealable aperture elements.

8. A method of transporting oocytes as described in claim 7, wherein said incubation element comprises a glass tube.

9. A method of transporting oocytes as described in claim 1, wherein said incubation conditions comprise an atmosphere of 5 percent carbon dioxide in air and a temperature of 39 degrees Centigrade within said incubation element.

10. A method of transporting oocytes as described in claim 1, further comprising the step of transferring said sperm cells to said straw containing said oocytes in said fertilization medium.

11. A method of transporting oocytes as described in claim 10, wherein said step of transferring sperm cells to said straw containing said oocytes comprises establishing a concentration of sperm cells in said fertilization medium of about 1 million to about 2 million per milliliter of fertilization medium.

12. A method of transporting oocytes as described in claim 2, further comprising the step of separating said sperm cells into enriched X-chromosome bearing and Y-chromosome populations.

13. A method of transporting oocytes as described in claim 12, further comprising the step of transferring separated sperm cells to said straw containing oocytes.

14. A method of transporting oocytes as described in claim 13, wherein said step of transferring separated sperm cells to said straw containing oocytes comprises establishing a concentration of said separated sperm cells in said fertilization medium of about 1 million to about 2 million per milliliter of fertilization medium.

15. A method of in vitro fertilization of oocytes, comprising the steps of:

a. collecting oocytes from a female non-human mammal;

b. entraining a plurality of oocytes in a fertilization medium containing both essential and non-essential amino acids;

c. inseminating said oocytes with sperm cells collected from a male mammal of the same species as said female mammal;

d. establishing inseminated oocytes in incubation conditions within an incubation element; and

e. fertilizing at least one of said oocytes.

16. A method of in vitro fertilization of oocytes as described in claim 15, wherein said fertilization medium is selected from the group consisting of chemically defined medium, SOF, or TALP supplemented with Eagles Medium.

17. A method of in vitro fertilization of oocytes as described in claim 15, wherein said female mammal is selected from the group consisting of primates, bovids, ovids, equids, swine, and dolphins.

18. A method of in vitro fertilization of oocytes as described in claim 15, wherein said non-essential amino acids have a concentration in said fertilization medium are futher supplemented with essential amino acids.

19. A method of in vitro fertilization of oocytes as described in claim 15, entraining a plurality of oocytes in a fertilization medium containing both essential and non-essential amino acids comprises entraining about 10 oocytes to about 15 oocytes in about 50 microliters of fertilization medium.

20. A method of in vitro fertilization of oocytes as described in claim 15, wherein said step of establishing oocytes in incubation conditions within said incubation element comprises an atmosphere of 5 percent carbon dioxide in air at a temperature between about 37 degrees Centigrade and about 41 degrees Centigrade for a duration of about 18 hours to about 20 hours.

21. A method of in vitro fertilization of oocytes as described in claim 15, wherein said step of inseminating said oocytes with sperm cells collected from a male mammal of the same species as said female mammal comprises establishing a concentration of said separated sperm cells in said fertilization medium of about 1 million to about 2 million per milliliter of fertilization medium.

22. A method of in vitro fertilization of oocytes as described in claim 15, wherein said sperm cells are separated on the basis of bearing an X-chromosome or bearing a Y-chromosome.