Methods for diagnosing human male infertility
First Claim
1. An in vitro method for screening for abnormal human sperm as part of a regimen for assessing human sperm fertilizing capacity comprising:
- obtaining a human sperm sample and isolating a number of sperm therefrom to provide a test sample;
incubating the test sample in a frog egg extract capable of supporting human sperm chromatin decondensation, DNA synthesis and chromatin recondensation;
monitoring the test sample for sperm chromatin decondensation and detecting an amount of test sperm decondensation; and
assessing the amount of human test sperm decondensation of the test sample and the amount of sperm chromatin decondensation of a human sperm control sample of proven fertile human sperm in the frog egg extract,wherein a test sample demonstrating about 80% or less decondensed sperm chromatin relative to the control sample amount of sperm chromatin decondensation provides a screen for abnormal human sperm.
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Abstract
A method for determining the capacity of a human sperm to fertilize a human egg is described by assessing sperm activation events in an in vitro assay using a non-mammalian egg extract, particularly a Xenopus laevis frog egg extract. Fertilizing capacity is assessed as a comparison of sperm decondensation, DNA synthesis and/or sperm recondensation as between a test sperm sample sperm and fertile sperm, such as a sperm sample from a proven fertile human male. The method employs results from the in vitro assay to also determine relative sufficiency or insufficiency of a sperm sample for fertilizing a human egg in human couples with a history of a diagnosed unexplained infertility from standard infertility diagnostic tests. The method may also be used to screen human sperm donors in human artificial insemination programs. A fixed-slide cytoprep sperm analysis of decondensed sperm chromatin, as between a sperm test sample and a sperm sample from a proven fertile human male, may also be used to confirm in vitro decondensation results of the infertility or fertility of a particular human male. A kit is also provided for testing male sperm samples for human egg fertilizing capacity.
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Citations
39 Claims
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1. An in vitro method for screening for abnormal human sperm as part of a regimen for assessing human sperm fertilizing capacity comprising:
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obtaining a human sperm sample and isolating a number of sperm therefrom to provide a test sample; incubating the test sample in a frog egg extract capable of supporting human sperm chromatin decondensation, DNA synthesis and chromatin recondensation; monitoring the test sample for sperm chromatin decondensation and detecting an amount of test sperm decondensation; and assessing the amount of human test sperm decondensation of the test sample and the amount of sperm chromatin decondensation of a human sperm control sample of proven fertile human sperm in the frog egg extract, wherein a test sample demonstrating about 80% or less decondensed sperm chromatin relative to the control sample amount of sperm chromatin decondensation provides a screen for abnormal human sperm. - View Dependent Claims (3, 4, 5, 6, 7, 8, 9, 10)
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2. A method for diagnosing infertility in a human male of unexplained infertility comprising:
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obtaining a sperm test sample from the human male of unexplained infertility; incubating the sperm test sample in a frog egg extract capable of supporting human sperm decondensation, DNA synthesis and chromatin recondensation; and comparing the amount of chromatin decondensation of the sperm test sample to the amount of chromatin decondensation of a sperm control sample from a human male of known fertility, wherein a sperm test sample demonstrating 80% or less decondensed sperm as compared to the sperm control sample is diagnostic of infertility in the human male of unexplained infertility.
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11. A method for screening human sperm as part of a regimen for selecting prospective fertile human sperm donors for in vitro fertilization of a human egg comprising:
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obtaining a sperm sample from a prospective human sperm donor to provide a test sperm sample; incubating the test sperm sample in a frog egg extract containing a detectable labeling compound; monitoring sperm chromatin decondensation of the test sperm sample; comparing the test sperm sample chromatin decondensation to a control sperm chromatin decondensation from a human male of proven fertility; and selecting a sperm test sample that demonstrates at least 80% of the control sperm chromatin decondensation as prospective donor sperm for in vitro fertilization of a human egg. - View Dependent Claims (12, 14, 15, 16)
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13. The method of clam 11 wherein the frog egg extract is a Xenopus laevis or Rana pipiens frog egg extract.
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17. A method for diagnosing infertility in a human male partner of an unexplained infertile human couple comprising the steps of:
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obtaining a sperm sample from the human male partner of the unexplained infertile human couple to provide a test sperm sample; incubating the test sperm sample in a frog egg extract capable of supporting human sperm chromatin decondensation to provide test sperm chromatin decondensation; determining the amount of test sperm chromatin decondensation; comparing the amount of the test sperm chromatin decondensation to an amount of sperm chromatin decondensation of control sperm of a proven fertile human male; and diagnosing infertility in the human male partner of the unexplained infertile human couple where the test sperm chromatin decondensation is 80% or less of the control sperm chromatin decondensation. - View Dependent Claims (18, 19, 20, 21)
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22. A toxicological screen for human sperm damage comprising:
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obtaining a sample of human sperm exposed to a toxic agent to provide a test sperm sample; incubating the test sperm sample in a frog egg extract capable of supporting human sperm chromatin decondensation to provide a test sperm chromatin decondensation; measuring the test sperm chromatin decondensation; and comparing the test sperm chromatin decondensation to a control sperm chromatin decondensation of proven fertile human male sperm, wherein a test sperm chromatin decondensation less than about 80% of the control sperm chromatin decondensation provides a toxicological screen for human sperm damage. - View Dependent Claims (23, 24)
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25. A method for screening varicocele human male patients as part of a regimen for screening patients for varicocelectomy comprising:
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monitoring chromatin decondensation in a sperm sample from the varicocele male and a proven fertile human sperm sample in a frog egg extract; and comparing decondensed nuclei in the varicocele sperm sample and the proven fertile human sperm sample, wherein a percentage of varicocele sperm decondensation less than 80% of the percentage of fertile sperm decondensation identifies a potential candidate for varicocelectomy. - View Dependent Claims (26, 27)
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28. A diagnostic kit for screening human males for infertility comprising:
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a carrier means adapted to receive at least four container means therein; a first container means containing a volume of dithiothreitol; a second container means containing a volume of lysolecithin; a third container means containing a non-mammalian egg extract capable of supporting fertile human sperm chromatin decondensation; a fourth container means containing a control sperm sample having a capacity for human sperm activation from a proven fertile human male; and at least two container means suitable for including therein a volume of up to 1 ml of a human sperm sample. - View Dependent Claims (29, 30, 31, 32, 33, 34, 35, 36)
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37. An in vitro method for screening human sperm for abnormalities as part of a regimen for assessing human sperm fertilizing capacity comprising:
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obtaining a human sperm sample and isolating a number of sperm therefrom to provide a test sample; incubating the test sample in a frog egg extract capable of supporting human sperm decondensation, DNA synthesis and sperm recondensation; monitoring the test sample for sperm chromatin decondensation, DNA synthesis and sperm chromatin recondensation and comparing sperm chromatin decondensation, DNA synthesis, and sperm chromatin recondensation of the test sample to a control sperm sample of a proven fertile human male, wherein a test sample demonstrating 80% or less of the chromatin decondensation, DNA synthesis and chromatin recondensation of the control sample provides a method for screening human sperm for abnormalities as part of a regimen for assessing human sperm fertilizing capacity. - View Dependent Claims (38, 39)
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Specification