A Noninvasive, Motility Independent, Sperm Sorting Method and Technology to Identify and Retrieve Individual Viable Nonmotile Sperm for Intracytoplasmic Sperm Injection
ABSTRACT For intracytoplasmic sperm injection in the absence of sperm motility it can be virtually impossible to distinguish viable from nonviable sperm. A reliable means to identify viable nonmotile sperm is needed and would likely improve the intracytoplasmic sperm injection success rate. Optoelectronic tweezers are a new technology that uses light induced dielectrophoresis fields to distinguish individual live cells from dead cells. We assessed the ability of optoelectronic tweezers to distinguish viable from nonviable individual nonmotile human sperm.
Fresh semen specimens from 6 healthy men were suspended in an isotonic sucrose/dextrose solution and incubated with 0.4% trypan blue dye (Sigma-Aldrich®). Within 15 minutes we randomly selected 5 motile and 50 nonmotile sperm, including 25 trypan negative, followed by 25 trypan positive sperm, under 200× magnification for optoelectronic tweezers assay. We recorded the individual sperm response (attraction or repulsion) to the optoelectronic tweezer field and trypan staining status.
From each subject 55 unwashed sperm were individually assayed for a total of 330. All motile sperm were attracted to optoelectronic tweezers. Of 150 trypan negative (viable) sperm 132 (88%) were attracted to the optoelectronic tweezer field with 0.88 sensitivity (95% CI 0.82-0.93) vs that of the trypan blue assay. All 150 trypan positive (nonviable) sperm were repulsed by or neutral to the optoelectronic tweezer field with 1.0 specificity (95% CI 0.98-1.00) vs that of the trypan blue assay. Type I error equaled 0 and overall assay agreement was 94%.
The optoelectronic tweezer assay can distinguish viable from nonviable nonmotile viable sperm with sensitivity comparable to that of the trypan blue assay and equal specificity. Optoelectronic tweezers are a promising means of selecting sperm for intracytoplasmic sperm injection.
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ABSTRACT: With the benefits of automation, sensitivity and precision, microfluidics has enabled complex and otherwise tedious experiments. Lately, lab-on-a-chip (LOC) has proven to be a useful tool for enhancing non-invasive assisted reproductive technology (ART). Non-invasive gamete and embryo assessment has largely been through periodic morpohological assessment using optical microscopy and early LOC ART was the same. As we realize that morphological assessment is a poor indication of gamete or embryo health, more advanced biophotonics has emerged in LOC ART to assay for metabolites or gamete separation via optoelectrical tweezers. Off-chip, even more advanced biophotonics with broad spectrum analysis of metabolites and secretomes has been developed that show even higher accuracy to predicting reproductive potential. The integration of broad spectrum metabolite analysis into LOC ART is an exciting future that merges automation and sensitivity with the already highly accurate and strong predictive power of biophotonics. (© 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim).Journal of Biophotonics 08/2012; 5(8-9). DOI:10.1002/jbio.201200041 · 3.86 Impact Factor
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ABSTRACT: Infertility studies are an important growing field, where new methods for the manipulation, enrichment and selection of sperm cells are required. Microfluidic techniques offer attractive advantages such as requirement of low sample volume and short processing times in the range of second or minutes. Presented here is the application of insulator-based dielectrophoresis (iDEP) for the enrichment and separation of mature and spermatogenic cells by employing a microchannel with cylindrical insulating structures with DC electric potentials in the range of 200-1500 V. The results demonstrated that iDEP has the potential to concentrate sperm cells and distinguish between mature and spermatogenic cells by exploiting the differences in shape which lead to differences in electric polarization. Viability assessments revealed that a significant percentage of the cells are viable after the dielectrophoretic treatment, opening the possibility for iDEP to be developed as a tool in infertility studies.Bioprocess and Biosystems Engineering 10/2012; 36(10). DOI:10.1007/s00449-012-0838-6 · 1.82 Impact Factor
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ABSTRACT: Introduction. Optical trapping is a laser-based method for probing the physiological and mechanical properties of cells in a noninvasive manner. As sperm motility is an important criterion for assessing the male fertility potential, this technique is used to study sperm cell motility behavior and rotational dynamics. Methods and Patients. An integrated optical system with near-infrared laser beam has been used to analyze rotational dynamics of live sperm cells from oligozoospermic and asthenozoospermic cases and compared with controls. Results. The linear, translational motion of the sperm is converted into rotational motion on being optically trapped, without causing any adverse effect on spermatozoa. The rotational speed of sperm cells from infertile men is observed to be significantly less as compared to controls. Conclusions. Distinguishing normal and abnormal sperm cells on the basis of beat frequency above 5.6 Hz may be an important step in modern reproductive biology to sort and select good quality spermatozoa. The application of laser-assisted technique in biology has the potential to be a valuable tool for assessment of sperm fertilization capacity for improving assisted reproductive technology.The Scientific World Journal 01/2014; 2014:154367. DOI:10.1155/2014/154367 · 1.73 Impact Factor