title: Use of laser tweezers to analyze sperm motility and mitochondrial membrane potential.
abstract: We combine laser tweezers with custom computer tracking software and robotics to analyze the motility [swimming speed, VCL (curvilinear velocity), and swimming force in terms of escape laser power (Pesc)] and energetics [mitochondrial membrane potential (MP)] of individual sperm. Domestic dog sperm are labeled with a cationic fluorescent probe, DiOC2(3), that reports the MP across the inner membrane of the mitochondria located in the sperm's midpiece. Individual sperm are tracked to calculate VCL. Pesc is measured by reducing the laser power after the sperm is trapped using laser tweezers until the sperm is capable of escaping the trap. The MP is measured every second over a 5-s interval during the tracking phase (sperm is swimming freely) and continuously during the trapping phase. The effect of the fluorescent probe on sperm motility is addressed. The sensitivity of the probe is measured by assessing the effects of a mitochondrial uncoupling agent (CCCP) on MP of free swimming sperm. The effects of prolonged exposed to the laser tweezers on VCL and MP are analyzed. The system's capabilities are demonstrated by measuring VCL, Pesc, and MP simultaneously for individual sperm. This combination of imaging tools is useful to quantitatively assess sperm quality and viability.
authors.affiliation: University of California, San Diego, Department of Electrical and Computer Engineering, 9500 Gilman Drive, La Jolla, California 92093, USA. jaclyn.nascimento@gmail.com
authors.forename: Jaclyn M
authors.initials: JM
authors.lastname: Nascimento
authors.forename: Linda Z
authors.initials: LZ
authors.lastname: Shi
authors.forename: Charlie
authors.initials: C
authors.lastname: Chandsawangbhuwana
authors.forename: James
authors.initials: J
authors.lastname: Tam
authors.forename: Barbara
authors.initials: B
authors.lastname: Durrant
authors.forename: Elliot L
authors.initials: EL
authors.lastname: Botvinick
authors.forename: Michael W
authors.initials: MW
authors.lastname: Berns
mesh_terms: D000818:Animals; D002469:Cell Separation / Q000295:instrumentation* / Q000379:methods; D002478:Cells, Cultured; D004285:Dogs; D004867:Equipment Design; D019544:Equipment Failure Analysis; D008297:Male; D053078:Membrane Potential, Mitochondrial / Q000502:physiology*; D052898:Optical Tweezers; D012371:Robotics / Q000295:instrumentation* / Q000379:methods; D013081:Sperm Motility / Q000502:physiology*; D013094:Spermatozoa / Q000502:physiology*