Separation systems of frozen-thawed spermatozoa into X-chromosome bearing and Y-chromosome bearing populations

  • US 7,771,921 B2
  • Filed: 09/28/2006
  • Issued: 08/10/2010
  • Est. Priority Date: 11/29/2000
  • Status: Active Grant
First Claim
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1. A process for preparing sperm cell samples, the process comprising:

  • collecting semen from a male mammal;

    freezing said semen to form frozen semen;

    thawing said frozen semen to form frozen-thawed semen;

    incubating sperm cells contained within said frozen-thawed semen in a concentration of Hoechst 33342 stain of greater than 40 micro-molar;

    establishing the temperature at which said sperm cells are incubated between about 30 degrees centigrade and about 40 degrees centigrade;

    adjusting a duration of time said sperm cells are incubated in said concentration of Hoechst 33342 stain;

    staining said sperm cells with sufficient uniformity to allow X-chromosome bearing sperm cells to be differentiated from Y-chromosome bearing sperm cells based upon the magnitude of fluorescence;

    determining the sex characteristic of a plurality of sperm cells contained within said frozen-thawed semen;

    separating said sperm cells according to the determination of their sex characteristic;

    isolating sperm cells separated according to the determination of their sex in a collection element;

    establishing a sample from said sperm cells isolated in said collection element;

    wherein fertilizing at least one egg of a female mammal with said sample at success levels of at least about 70% of success with spermatozoa that have not been separated and/or frozen is achieved.

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