Sdad assay and uses thereof

a technology of assay and sdad, applied in the field of assay, can solve the problem that the predictive capacity of screening methods is relatively low, and achieve the effect of low probability of successful art attempts at pregnancy

Inactive Publication Date: 2009-02-26
BROWN DAVID
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0007]In some embodiments, a screening method is provided that is useful in identifying male sperm donors who will have a low probability of successful ART attempts at pregnancy (success being defined as live birth), such as when using ICSI. Other sperm screening assays, such as the sperm DNA decondensation (SDD) Test, the sperm penetration assay (SPA), and the sperm chromatin structure assay (SCSA) have all been found to not be predictive of ICSI live birth outcome. Thus, in some aspect, the present disclosure describes a screening method that can predict ICSI outcome. In some embodiments, this screening method comprises obtaining a se

Problems solved by technology

The screening method has relatively low predictive capacity for determining a couple's success, success being defined as live birth, when sperm from the male partner is used in an ICSI attempt at pre

Method used

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  • Sdad assay and uses thereof
  • Sdad assay and uses thereof
  • Sdad assay and uses thereof

Examples

Experimental program
Comparison scheme
Effect test

example 1

Materials and Methods

[0066]The present example provides a detailed description of some of the materials and methods employed in the examples that follow.

[0067]Materials and Methods:

[0068]Chemicals:

[0069]Lipid peroxidation of the sperm cellular membrane was determined using the Oxford Biomedical Research Kit Fr 12.

[0070]Autoradiography Emulsion, Type NTB2; and photographic developer D-19 by KODAK (Eastman Kodak, Rochester, N.Y.). All the chemicals were purchased from Sigma Chemical Company (St Louis, Mo., USA).

[0071]Experimental Design:

[0072]In vitro oxidative stress exposure; human sperm from a fertile male were exposed to increasing hydrogen peroxide H2O2 concentrations (0, 10, 50 and 100 μM), and incubated at 37° C., for 15 min and 1 hr.

[0073]Lipid peroxidation (LPO) measurements and HSAA responses were analyzed in each treatment to determine the effect(s) H2O2 concentration- and time exposure on human sperm activation responses.

[0074]Hydrogen Peroxide Dose-Response:

[0075]Sperm Pr...

example 2

Use of the HSAA and the SDAD Test in Identifying Reactive Oxygen Species (ROS) Damage

[0085]The present example is provided to demonstrate the utility of the HSAA and the 5-minute time point SDAD Test, an addition to the HSAA, in the identification of damage resulting from exposure to ROS.

[0086]The HSAA as described in Example 1 was used in the present study.

[0087]Experimental Design:

[0088]In vitro oxidative stress exposure; human sperm from a fertile male were exposed to increasing hydrogen peroxide (H2O2) concentrations (0, 10, 50 and 100 μM), and incubated at 37° C., for 15 min and 1 hr.

[0089]Lipid peroxidation (LPO) measurements and HSAA responses were analyzed in each treatment to determine the effect(s) of H2O2 concentration and time exposure on human sperm activation responses.

[0090]Hydrogen Peroxide Dose-Response:

[0091]Sperm Preparation and Oxidative Treatments—

[0092]Semen ejaculate was collected in a sterile jar by masturbation after 3-5 days of sexual abstinence from fertil...

example 3

Sperm DNA Decondensation (SDD) and Sperm Penetration Assay (SPA) with Gradient Preparation Are Not Predictive of Pregnancy Outcome Using In Vitro Fertilization (IVF) with Intracytoplasmic Sperm Injection (ICSI) Cycles

[0145]Several sperm structure tests that detect compromised DNA integrity have been shown to have poor predictive capacity for successful ICSI cycles. This is believed to be the result of improved selection of the best sperm for use in ICSI, by use of a density gradient. Two different sperm function tests, the SDD Test and the SPA, are examined for capacity to predict success in ICSI cycles.

[0146]The SDD Test measures the fraction of cells fully decondensed after a 15 minute incubation in frog egg extract, measuring delayed decondensation, while the SPA measures the sperm capacitation index (SCI), the average number of sperm penetrations per zona-free hamster oocyte. The objective of this study was to review the pregnancy outcome in ICSI cycles in relation to the SPA an...

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Abstract

Methods for screening human sperm using a sperm DNA accelerated decondensation (SDAD) Test, readable at a 5 minute time point, are disclosed. The method is useful in the screening and clinical management for a reproductively challenged human couple. The methods disclosed are highly predictive of ICSI outcome. Methods for screening human sperm using a sperm DNA decondensation (SDD) Test, readable at a 15 minute time point, are disclosed. This test is highly predictive of IUI and IVF outcome. Also dislosed are methods for selecting an appropriate assisted reproductive technique (ART) for a reproductively challenged couple. These methods provide a decision tree that employs a patient's sperm test results in a sperm DNA accelerated decondensation (SDAD) Test, and sperm DNA decondensation (SDD) Test, wherein an appropriate assisted technique may be selected from intrauterine insemination (IUI), IVF or ICSI. Also, methods are described that will identify males who may benefit from anti-oxidant therapy and/or a varicocelectomy before using their sperm in ART. An automated scoring method is also disclosed for assessing sperm activity after a 5-minute incubation in egg extract (SDAD Test), and after a 15-minute incubation in egg extract (SDD Test). Finally, an automated sperm processing protocol is also disclosed for the rapid preparation of sperm for analysis in both the SDD and SDAD Tests.

Description

PRIORITY CLAIM[0001]This application is a non-provisional of, claims priority to and the benefit of U.S. Provisional Patent Application Ser. No. 60 / 957,117, filed on Aug. 21, 2007, the entire contents of which are incorporated herein by reference.BACKGROUND OF INVENTION[0002]The process of fertilization involves a series of events including: a) sperm binding to, and then penetration of the oocyte zona pellucida via the acrosome reaction, b) sperm fusion with the oocyte plasma membrane, c) oocyte activation (release of the cortical granules to prevent multiple sperm from fertilizing the egg), d) sperm activation and pronucleous formation (Abou-Haila A. et al. 2000: Furlong, L. et al. 2005).[0003]Sperm activation initiates post-fertilization upon entry into the ooplasm where the compacted sperm DNA undergoes many alterations as it develops into a male pronucleus. During the first phase of sperm activation or decondensation, the chromatin condensation acquired during spermiogenesis is ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
CPCG01N2800/367G01N33/5091A61D19/02C12Q1/68
Inventor BROWN, DAVID
Owner BROWN DAVID
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