Changes in plasma membrane and acrosome integrity of frozen-thawed bovine spermatozoa during a 4 h incubation as measured by multicolor flow cytometry

Szabolcs Nagy, Triin Hallap, Anders Johannisson, Heriberto Rodriguez-Martinez

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37 Citations (Scopus)


Cryopreservation of bull semen is sub-optimal, causing cell death of a majority of spermatozoa. Even the surviving cells are affected post-thaw, either structurally or functionally. The aim of this study was to investigate the sequence of events that take place when sperm plasma membrane and acrosome deteriorate during a 4h incubation period post-thaw, with special attention paid to the acrosome status of dying cells. Frozen-thawed semen of six AI dairy bulls was used. Three straws per batch were pooled and incubated at 37°C. Sub-samples were taken at 30min intervals and stained with SYBR 14, propidium iodide (PI) and phycoerythrin-conjugated peanut agglutinin (PE-PNA). Plasma membrane and acrosome integrity were measured by flow cytometry. The experiment was repeated three times. Immediately after thawing, only 3.45% of the dying cells showed acrosomal exocytosis. This number increased dramatically during incubation, reaching 67% after 4h. Within the intact cell population, the overall decrease in viability and acrosome integrity was kept at five percentage points. Flow cytometry and the triple fluorochrome combination presented a detailed picture of the time course in plasma membrane and acrosome deterioration of frozen-thawed bull semen. The results are expected to be useful for monitoring new cryopreservation protocols.

Original languageEnglish
Pages (from-to)225-235
Number of pages11
JournalAnimal Reproduction Science
Issue number3-4
Publication statusPublished - Feb 2004



  • Acrosome
  • Bovine sperm
  • Cryopreservation
  • Flow cytometry
  • Plasma membrane

ASJC Scopus subject areas

  • Food Animals
  • Animal Science and Zoology
  • Endocrinology

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