Devices and methods for quantification of male fertility

Patent metadata

Field	Value
Publication number	US-10852298-B2
Application number	US-201615775612-A
Country	US
Kind code	B2
Filing date	Nov 18, 2016
Priority date	Nov 20, 2015
Publication date	Dec 1, 2020
Grant date	Dec 1, 2020

Abstract

Disclosed herein is a low cost and rapid microfluidic based method and test device for quantifying male fertility potential. The device can simultaneously measure three critical semen parameters rapidly, namely live sperm concentration, motile sperm concentration, and sperm motility. The device includes a transparent substrate and a top sheet with two holes therethrough and an intermediate sheet sandwiched between the substrate and the top sheet. The wells formed by holes form a concentration measuring well (C) and a motility well (M) formed by the top sheet with these two holes bonded to the intermediate sheet. A colorimetric agent is located on the top surface of the intermediate sheet at the bottom of each well which changes color when in contact with sperm. In the motility well a porous membrane is located on top of the colorimetric agent and a liquid buffer may be placed on the top surface of the porous membrane. Applying part of a sperm sample to the C well results in direct contact of any live sperm with the colorimetric agent causing a color change, applying part of the sperm sample to the M well results in live sperm with sufficient motility to swim vertically down through the liquid buffer and through the porous membrane to the colorimetric agent. Evaluating the intensities of the color change of the colorimetric agents before and after contact with the sample gives a measure of total concentration of live sperm and motile sperm from which sperm motility is calculated.

First claim

Therefore what is claimed is:

1. A device for quantifying fertility potential from a semen sample by calculating total concentration of live sperm, total concentration of live motile sperm and sperm motility in the sample as compared with reference values defined by the World health Organization, comprising:
a transparent or semitransparent substrate layer;
a first layer comprised of a hydrophilic porous material secured on top of the substrate layer, the first layer having first and second reaction spots spaced from each other and each reaction spot containing a colorimetric agent selected to undergo reaction with sperm such that upon reaction with sperm a color change occurs;
a porous membrane covering said second reaction spot, said membrane having pores in a size range large enough to allow sperm to swim through the membrane and small enough to prevent the sperm in the semen sample to flow through the membrane;
a second layer comprised of a hydrophilic porous material secured on top of the first layer, said second layer including first and second holes being located with respect to each other such that when the second layer is affixed to a top surface of the first layer said first and second holes are aligned with the first and second reaction spots; and
wherein upon application of a first portion of a semen sample directly to said first reaction spot through said hole in said second layer a color change occurs when live sperm is present and the intensity of reacted colorimetric agent is measured using a software package to calculate total concentration of live sperm, and upon application of a second portion of the semen sample through the second hole onto the membrane any change in color indicates motile sperm is present in the second portion that traversed through the porous membrane to contact the second reaction spot and the intensity of reacted colorimetric agent located in the second reaction spot is used to calculate total concentration of live motile sperm, and the ratio of total live motile sperm concentration to total live sperm concentration is used to calculate sperm motility.

2. The device according to claim 1 , including a third spot located on said second layer spaced from the first and second reactions spots, said third spot having a colorimetric agent coated thereon, said first layer not including a hole in registration with said third spot.

3. The device according to claim 1 , including a buffer solution formulated to have a viscosity to simulate in vivo fluid in the female tract, said buffer solution configured to be applied to a top surface of the porous membrane prior to application of a semen sample thereto such that in operation any live sperm must traverse through the buffer solution and then through the porous membrane.

4. The device according to claim 3 , including an integrated sealed buffer reservoir having a removable seal configured to be released by a user onto the top surface of the porous membrane, said buffer chamber being in flow communication with said surface of the porous membrane.

5. The device according to claim 4 , including wherein the integrated sealed buffer reservoir is in flow communication with said surface of the porous membrane via a channel to the membrane from the buffer reservoir.

6. The device according to claim 1 , wherein said porous membrane is hydrated, and including a separate detachable cover covering the top surface of the membrane for maintaining its moisture content while maintaining the rest of device dry.

7. The device according to claim 1 , wherein the porous membrane is selected such that it is saturated with the second portion of the semen sample to facilitate flow through the porous membrane.

8. The device according to claim 1 including an analysis system and a software package, the software package configured to be programmed into an analysis system, said analysis system including a color sensor configured to sense a color of the colorimetric agents in the first and second reaction spots through a bottom surface of said transparent or semitransparent substrate layer.

9. The device according to claim 8 wherein said software package is programmed with instructions to calculate
a total concentration of live sperm in the first portion of semen located on the first reaction spot based on an intensity of reacted colorimetric agent in said first reaction spot, and
a total concentration of live motile sperm in the second portion of semen that traversed through said liquid buffer and said porous membrane to said second reaction spot based on an intensity of reacted colorimetric agent in said second reaction spot.

10. The device according to claim 9 wherein said software package is programmed with instructions to calculate sperm motility based on the total concentration of live sperm and total concentration motile sperm.

11. The device according to claim 8 wherein said analysis system includes a visual display, and wherein said software package is programmed with instructions to visually display said total concentration live sperm and total concentration of motile sperm, and sperm motility.

12. The device according to claim 8 wherein the analysis system is a computer system interfaced with a camera programmed with said software package, said computer system having a visual display.

13. The device according to claim 8 wherein the analysis system is a smart phone having a built-in camera and said software package is a smart phone application loaded onto said smart phone.

14. The device according to claim 1 wherein the first and second layers are made of paper based hydrophilic porous materials.

15. The device according to claim 1 wherein the first and second layers are made of polymer based hydrophilic porous material.

16. The device according to claim 1 configured for testing human sperm, and wherein the membrane has pores in a size range from about 5 to about 12 microns.

17. The device according to claim 1 wherein the membrane has pores in a size range from about 4 to about 14 microns.

18. A method for quantifying fertility potential from a semen sample by calculating total concentration of live sperm, total concentration of live motile sperm and sperm motility in the sample as compared with reference values defined by the World health Organization, comprising:
a) applying a first portion of a semen sample onto a colorimetric agent localized on a first reaction area on a support structure;
b) applying a second portion of the semen sample onto a second reaction area on the support structure, the second reaction area including a colorimetric agent localized on a second reaction area on the support structure and a porous membrane located on top of the colorimetric agent and a liquid buffer having sufficient viscosity to simulate in vivo fluid in the female tract located on top of the porous membrane such that motile sperm in the second portion of the semen sample must pass first through the liquid buffer and then the porous membrane to reach the colorimetric agent located on the second reaction area of the support structure, the porous membrane having pores in a size range large enough to allow sperm to swim through the membrane and small enough to prevent the sperm in the semen sample to flow through the membrane; and
c) observing any color change in the colorimetric agents localized on the first and second reaction areas, and when color changes are present, comparing an intensity change on the first reaction area to an intensity of unreacted colorimetric agent and comparing an intensity change

Assignees

• Governing Council Univ Toronto

Inventors

• Nosrati Reza
• Gong Max M
• Sinton David

Classifications

• G01N33/48707

  by electrical means (G01N33/49, G01N33/493 take precedence) · CPC title

• C12N5/0612

  sorting of gametes, e.g. according to sex or motility · CPC title

• G01N2015/0681

  Purposely modifying particles, e.g. humidifying for growing · CPC title

• G01N30/00

  Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography {or field flow fractionation} (G01N3/00, G01N5/00, G01N7/00, G01N9/00, G01N11/00, G01N13/00, G01N15/00, G01N17/00, G01N19/00, G01N21/00, G01N22/00, G01N23/00, G01N24/00, G01N25/00, G01N27/00, G01N29/00 take precedence) · CPC title

• C12N5/061

  Sperm cells, spermatogonia · CPC title