Fractionation of sperm

• US 4,009,260 A
• Filed: 12/11/1974
• Issued: 02/22/1977
• Est. Priority Date: 04/19/1973
• Status: Expired due to Term

• Current Assignee: Gametrics Limited
• Original Assignee: Schering Ag (Merck & Co., Inc.)
• Inventors: Ericsson, Ronald J.
• Primary Examiner(s): Rosen, Sam
• Application Number:
  US05/531,728
• Time in Patent Office: 804 Days
• Field of Search: 195/1.8, 424/105
• US Class Current: 424/561
• CPC Class Codes: A61B 17/43 for artificial insemination
• CPC Class Codes: A61D 19/00 Instruments or methods for reproduction or fertilisation
• CPC Class Codes: A61K 35/52 Sperm; Prostate; Seminal fluid; Leydig cells of testes

Abstract

Motile sperm are separated from immotile and abnormal sperm of mammalian semen by maintaining, at a temperature at which the motile sperm are motile, a layer of at least the sperm portion of the semen in interfacial contact with a discrete lower layer of an aqueous contacting medium physiologically acceptable to the sperm in which the motile sperm have a lower migration rate than in the upper layer, until the motile sperm of the semen have migrated at a preferential rate to the contacting medium, thereby providing a sperm fraction consisting essentially of normal motile sperm with enhanced Y-sperm content. Repeating the process with a second contacting medium physiologically acceptable to the sperm which further retards the migration rate of the motile sperm increases substantially the Y- to X-sperm ratio of the sperms which migrate thereto. The resulting sperm fraction, because of its enhanced Y-sperm content, can be used for artificial insemination to increase the incidence of male offspring. The first step alone can be used to improve the quality of semen having an abnormally low normal motile sperm content, thereby increasing the chances of a successful insemination.

Claims

1. A process for increasing the proportion of Y-sperm in mammalian sperm, which comprises the steps of a) maintaining at least the sperm portion of progressively motile sperm-containing semen, either as such or suspended in an aqueous suspending vehicle physiologically acceptable to the sperm, as an upper layer in vertical interfacial contact, at a temperature at which the motile sperm are motile, with a lower discrete layer of a first aqueous contacting medium physiologically acceptable to the sperm and in which the motile sperm migrate downwardly at a slower rate than in the upper layer, until a portion only of the motile sperm of the semen having migrated downwardly into the contacting medium, thereby producing a contacting medium containing a higher proportion of Y to X sperm than in the starting sperm, and b) repeating the step at least once, employing in the upper layer the motile sperm which have migrated to the first aqueous contacting medium and as the lower layer a second aqueous contacting medium physiologically acceptable to the sperm and in which the motile sperm migrate downwardly at a slower rate than in the first contacting medium.

2. A process according to claim 1 wherein the sperm is human sperm.

3. A process according to claim 1 wherein the sperm is bovine sperm.

4. A process according to claim 1 wherein the sperm is porcine sperm.

5. A process according to claim 1 wherein the starting sperm is separate from the other components of the semen.

6. A process according to claim 1 wherein the starting sperm is suspended in an aqueous suspending vehicle physiologically acceptable to the sperm which does not retard the downward migration rate of the motile sperm.

7. A process according to claim 6 wherein the starting sperm is suspended in Tyrode solution.

8. A process according to claim 1 wherein the contacting medium comprises a serum albumin, ovalbumin or a serum globulin.

9. A process according to claim 8 wherein the contacting medium is a 3-25% solution of bovine serum albumin in Tyrode solution.

10. A process according to claim 1 wherein in each step the sperm layer is contacted at about room temperature with the contacting medium for about 0.5 to 10 hours.

11. A process according to claim 10 wherein the sperm is contacted with the first contacting medium for about 1 to 3 hours.

12. A process according to claim 1 wherein the second contacting medium is positioned below the first contacting medium while the latter is positioned below the starting sperm layer.

13. A process according to claim 12 wherein the starting sperm is separate from the other components of semen and is suspended in an aqueous suspending vehicle physiologically acceptable to the sperm which does not retard the downward migration rate of the sperm.

14. A process according to claim 13 wherein the sperm is human sperm.

15. A process according to claim 12 wherein the starting sperm layer is separated from the first contacting medium before the second contacting step is completed.

16. A process according to claim 1 wherein the sperm which migrates to the first contacting medium is separated therefrom and suspended in an aqueous suspending vehicle physiologically acceptable to the sperm which does not retard the downward migration rate of the motile sperm before being contacted with the second contacting medium.

17. A process according to claim 16 wherein the starting sperm is separate from the other components of semen and is suspended in an aqueous suspending vehicle physiologically acceptable to the sperm which does not ratard the downward migration rate of the sperm.

18. A process according to claim 17 wherein the sperm is human sperm.

19. A process according to claim 1 wherein after motile sperms have migrated into the first contacting medium the latter is separated from the starting sperm layer and thereafter contacted with the second contacting medium.

20. A process according to claim 19 wherein the starting sperm is separate from the other components of semen and is suspended in an aqueous suspending vehicle physiologically acceptable to the sperm which does not retard the downward migration rate of the sperm.

21. A process according to claim 20 wherein the sperm is human sperm.

34. A method for increasing the incidence of males in mammalian offspring which comprises artificially inseminating a fertile female with Y-sperm enriched sperms of the same species of mammal obtained according to the process of claim 1.

35. A method according to claim 34 wherein the mammal is human.

36. A method according to claim 34 wherein the mammal is bovine.

37. A method according to claim 34 wherein the mammal is porcine.

22. A process for increasing the proportion of motile to immotile sperm in mammalian sperm, which comprises the steps of a) separating the sperm from the other components of the semen;

b) suspending the separated sperm portion of the semen in an aqueous suspending vehicle physiologically acceptable to the sperm and which does not substantially retard the downward migration rate of motile sperm;

c) maintaining the sperm-containing suspending vehicle as an upper layer in interfacial contact, at a temperature at which the motile sperm are motile, with a lower discrete layer of an aqueous contacting medium which is physiologically acceptable to the sperm and in which the motile sperm migrate downwardly at a slower rate than in the suspending vehicle, until at least a portion of the motile sperm of the separated sperm have migrated downwardly into the contacting medium, thereby producing a sperm fraction having a higher proportion of motile sperm than in the starting sperm; and

d) thereafter separating the contacting medium, containing the motile sperm enhanced fraction of the starting sperm, from the suspending vehicle layer.

23. A process according to claim 22 wherein the sperm is human sperm.

24. A process according to claim 22 wherein the sperm is bovine sperm.

25. A process according to claim 22 wherein the sperm is equine sperm.

26. A process according to claim 22 wherein the motile sperm which migrate to the contacting medium are thereafter separated therefrom.

27. A process according to claim 22 wherein the starting sperm is suspended in Tyrode solution.

28. A process according to claim 22 wherein the contacting medium comprises a serum albumin, ovalbumin or a serum globulin.

29. A process according to claim 28 wherein the contacting medium is a 3-25% solution of bovine serum albumin in Tyrode solution.

30. In a method for artificially inseminating a mammalian female, the improvement which comprises employing motile sperm enriched sperm obtained according to the process of claim 22.

31. A method according to claim 30 wherein the mammal is a human.

32. A method according to claim 30 wherein the mammal is a bovine.

33. A method according to claim 30 wherein the mammal is an equine.