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Evaluation of acrosome integrity

CN Chiara Nerozzi
SR Sandra Recuero
GG Giovanna Galeati
DB Diego Bucci
MS Marcella Spinaci
MY Marc Yeste
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Acrosome integrity was determined through staining with the lectin from Arachis hypogaea (PNA) conjugated with FITC (fluorescein isothiocyanate; PNA-FITC) and PI. Five hundred μL of each sperm sample was incubated with 0.5 μL PNA-FITC (final concentration: 1.25 mg/mL) and 2.5 μL PI (final concentration: 12 μM) for 10 min at 38 °C in darkness. Spermatozoa were then evaluated with the flow cytometer and two categories were distinguished: (a) viable spermatozoa with intact acrosome and plasma membrane (PNA-FITC−/PI−), and (b) spermatozoa that had damaged their plasma membrane and/or their acrosome, which included PNA-FITC+/PI−, PNA-FITC+/PI+, PNA-FITC−/PI+ populations. Debris, non-stained particles found in SYBR14/PI staining (i.e. SYBR14−/PI−) were subtracted from the PNA-FITC-/PI- population and the other percentages were recalculated.

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