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The MISS may decrease risks24 associated with intracytoplasmic sperm injection by isolating viable sperm using a mild, biomimetic sorting mechanism based on sperm motility; it avoids centrifugation and sample compaction, Florence, L

Passively Driven Integrated Microfluidic System for Separation of Motile Sperm

ANALYTICAL CHEMISTRY, no. 7 (2003): 1671-1675

Cited by: 295|Views17
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Abstract

This paper describes a self-contained integrated microfluidic system that can separate motile sperm from small samples that are difficult to handle using conventional sperm-sorting techniques. The device isolates motile sperm from nonmotile sperm and other cellular debris, based on the ability of motile sperm to cross streamlines in a lam...More

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Introduction
  • The device fulfills a need in clinical settings where small amounts of sperm need to be sorted.
  • It opens the way for convenient bioassays based on sperm motility including at-home motile sperm tests.
  • While theoretically only a single sperm is needed per harvested ooctye, the practicality of processing, isolating, and locating the most viable sperm is challenging, or in some cases impossible, with current techniques such as centrifugation and swim-up processes..
  • Isolation of viable sperm from samples with severely low sperm counts or from small amounts of cryopreserved spermsfor example, from patients that have preserved sperm prior to undergoing chemotherapysrequires new efficient technologies
Highlights
  • This paper describes a self-contained integrated microfluidic system that can separate motile sperm from small samples that are difficult to handle using conventional sperm-sorting techniques
  • The MISS uses a sorting system where nonmotile sperm flow along their initial streamlines and exit one outlet whereas motile sperm can deviate from their initial streamlines and exit through a different outlet (Figure 2)
  • This sorting mechanism is related to the “filtering” mechanism used in the H-filter where rapidly diffusing small molecules exit through a different outlet from larger molecules and particles that diffuse more slowly
  • The MISS increases the efficiency of sperm isolation from samples with few motile sperm that are difficult or impossible to process using conventional methods
  • The MISS may decrease risks24 associated with intracytoplasmic sperm injection (ICSI) by isolating viable sperm using a mild, biomimetic sorting mechanism based on sperm motility; it avoids centrifugation and sample compaction, (20) Florence, L
Methods
  • Sorting tests were performed using washed semen samples.
  • 60 μL of processing medium was added to the media inlet reservoir, 50 μL of a washed semen sample to the sample inlet reservoir, and 2 μL of medium to each of the outlet reservoirs.
  • For visualization of membrane-compromised sperm, which generally corresponds to nonmotile sperm, 3 μL of PI (Molecular Probes, www.probes.com, 60 mM dissolved in processing medium) was added to sperm samples prior to sorting.
  • An inverted microscope (Nikon TE 300, www.nikon-usa.com) with a CCD camera (Hamamatsu ORCA-100, www.hamamatsu.com) was used to capture images and record movies
Results
  • The MISS uses a sorting system where nonmotile sperm flow along their initial streamlines and exit one outlet whereas motile sperm can deviate from their initial streamlines and exit through a different outlet (Figure 2).
  • The difference between the two devices is that the MISS takes advantage of active movement of cells whereas the H-filter takes advantage of passive diffusion of particles
  • These types of sorting are possible because, in small channels, multiple laminar streams can flow parallel to each other with no turbulent mixing at the interface between the streams..
Conclusion
  • New single-cell-based treatments such as ICSI require new technologies for cellular manipulations in order to make procedures more efficient and safe. The MISS increases the efficiency of sperm isolation from samples with few motile sperm that are difficult or impossible to process using conventional methods.
  • New single-cell-based treatments such as ICSI require new technologies for cellular manipulations in order to make procedures more efficient and safe..
  • The MISS increases the efficiency of sperm isolation from samples with few motile sperm that are difficult or impossible to process using conventional methods.
  • The MISS may decrease risks associated with ICSI by isolating viable sperm using a mild, biomimetic sorting mechanism based on sperm motility; it avoids centrifugation and sample compaction,.
Summary
  • Introduction:

    The device fulfills a need in clinical settings where small amounts of sperm need to be sorted.
  • It opens the way for convenient bioassays based on sperm motility including at-home motile sperm tests.
  • While theoretically only a single sperm is needed per harvested ooctye, the practicality of processing, isolating, and locating the most viable sperm is challenging, or in some cases impossible, with current techniques such as centrifugation and swim-up processes..
  • Isolation of viable sperm from samples with severely low sperm counts or from small amounts of cryopreserved spermsfor example, from patients that have preserved sperm prior to undergoing chemotherapysrequires new efficient technologies
  • Methods:

    Sorting tests were performed using washed semen samples.
  • 60 μL of processing medium was added to the media inlet reservoir, 50 μL of a washed semen sample to the sample inlet reservoir, and 2 μL of medium to each of the outlet reservoirs.
  • For visualization of membrane-compromised sperm, which generally corresponds to nonmotile sperm, 3 μL of PI (Molecular Probes, www.probes.com, 60 mM dissolved in processing medium) was added to sperm samples prior to sorting.
  • An inverted microscope (Nikon TE 300, www.nikon-usa.com) with a CCD camera (Hamamatsu ORCA-100, www.hamamatsu.com) was used to capture images and record movies
  • Results:

    The MISS uses a sorting system where nonmotile sperm flow along their initial streamlines and exit one outlet whereas motile sperm can deviate from their initial streamlines and exit through a different outlet (Figure 2).
  • The difference between the two devices is that the MISS takes advantage of active movement of cells whereas the H-filter takes advantage of passive diffusion of particles
  • These types of sorting are possible because, in small channels, multiple laminar streams can flow parallel to each other with no turbulent mixing at the interface between the streams..
  • Conclusion:

    New single-cell-based treatments such as ICSI require new technologies for cellular manipulations in order to make procedures more efficient and safe. The MISS increases the efficiency of sperm isolation from samples with few motile sperm that are difficult or impossible to process using conventional methods.
  • New single-cell-based treatments such as ICSI require new technologies for cellular manipulations in order to make procedures more efficient and safe..
  • The MISS increases the efficiency of sperm isolation from samples with few motile sperm that are difficult or impossible to process using conventional methods.
  • The MISS may decrease risks associated with ICSI by isolating viable sperm using a mild, biomimetic sorting mechanism based on sperm motility; it avoids centrifugation and sample compaction,.
Funding
  • We thank the Whitaker Foundation, NICHD, and the Engineering Technology Development Fund for financial support. SUPPORTING INFORMATION AVAILABLE Three movies (one showing separation of motile from nonmotile sperm in the microfluidic channel, one showing flow of propidium iodide-stained sperm viewed under green excitation light, and another showing flow of propidium iodide-stained sperm along with nonstained motile sperm viewed under phase contrast illumination)
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