Hum. Reprod. Advance Access originally published online on March 2, 2007
Human Reproduction 2007 22(5):1405-1412; doi:10.1093/humrep/dem015
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Molecular glass wool filtration as a new tool for sperm preparation
1 Department of Dermatology/Andrology Unit, University of Leipzig, Leipzig, Germany 2 Department of Dermatology and Andrology, Justus Liebig University, Giessen, Germany
3 To whom correspondence should be addressed at: Department of Dermatology, Training Center of the European Academy of Andrology, University of Leipzig, Philipp-Rosenthal-Strasse 23-25, 04103 Leipzig, Germany. Tel.: +49 341 9718611; Fax: +49 341 9718619; E-mail: uwe.paasch{at}medizin.uni-leipzig.de
BACKGROUND: Magnetic-activated cell sorting (MACS) using annexin V-conjugated microbeads in a liquid phase eliminates apoptotic spermatozoa based on the externalization of phosphatidylserine (EPS) residues. The procedure allows the enrichment of a sperm population free of apoptosis markers, giving higher fertilization potential. Our aim was to determine if the annexin V binding principle can be transferred onto a glass wool filter system in order to produce a solid phase filter.
METHODS: Semen samples (n = 42) were subjected to a molecular glass wool filter system using glass surfaces coated with annexin V and compared with aliquots separated by conventional glass wool, as well as with annexin V-MACS. The extent of apoptosis was assessed by measuring levels of activated caspase 3 using fluorescein-labelled inhibitors of caspase, alterations in mitochondrial membrane potential (MMP) using a lipophilic cationic dye, and EPS using a fluorescein isothiocyanate-coupled monoclonal antibody.
RESULTS: Annexin V-negative sperm filtered out by the newly developed molecular glass wool filtration (GWF) system displayed superior quality in terms of high MMP integrity, as well as, to a small extent, caspase 3 activation and EPS.
CONCLUSIONS: The effect of traditional GWF can be further improved by combination with annexin V binding. This newly developed solid phase molecular filter system has been proven to enrich spermatozoa free of apoptosis markers to the same extent as the annexin V magnetic separation technique. The selection of spermatozoa free of apoptosis markers by molecular glass wool filters may enhance the results of IVF.
Key words: annexin V/apoptosis marker/glass wool/IVF/sperm
Submitted on November 21, 2006; accepted on January 9, 2007.