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Start Over Descriptor "−80°C freezer" Topic sperm cryopreservation
2 results on '"−80°C freezer"'

1. The impact of five years storage/biobanking at −80°C on mouse spermatozoa fertility, physiology, and function.

Author

Raspa, Marcello, Putti, Sabrina, Paoletti, Renata, Barboni, Barbara, Ramal‐Sanchez, Marina, Lanuti, Paola, Marchisio, Marco, D'Atri, Mario, Ortolani, Claudio, Papa, Stefano, Valbonetti, Luca, Bernabo, Nicola, and Scavizzi, Ferdinando

Subjects
*SPERMATOZOA, *FERTILIZATION in vitro, *PHYSIOLOGY, *FERTILITY, *INTRACELLULAR calcium
Abstract

Background: We previously demonstrated how mouse spermatozoa can be efficiently stored for two years in a −80°C freezer, maintaining their ability to fertilize mouse eggs. Objectives: The main objective here was to evaluate the effects of five years at −80°C compared to liquid nitrogen storage (LN2, control condition) on mouse sperm viability, physiological parameters, and fertilization capacity. Materials and methods: Three different strains were used: C57BL/6N, C57BL/6J and CD1. Flow cytometry experiments were performed to analyze sperm viability (SYBR‐14 + Propidium Iodide +Hoechst33342), the intracellular calcium concentration (Fluo 3‐AM), the membrane lipid disorder (Merocyanine 540), and the mitochondrial activity (MitoTracker Red) in live spermatozoa. The in vitro fertilization (IVF) was used to evaluate the sperm fertilizing ability. Results: Flow cytometry analysis showed that the percentage of live cells are reduced in B6N and B6J, but not in CD1 mice. However, in the live population no differences in terms of intracellular calcium concentration, membrane lipid disorder, and mitochondrial activity were reported when comparing both biobanking methods. Spermatozoa stored at −80°C for 5 years successfully fertilized the eggs and developed mouse embryo normally both in culture and in vivo, generating live pups with no differences compared to control samples stored in LN2. Discussion: Long‐term mouse sperm storage at −80°C (five years) could be considered an ideal alternative to the most common LN2 approach, giving economical and logistic advantages. Moreover, the precise information originated from the flow cytometry analysis stands up this technique as an optimal strategy to evaluate the sperm quality and ranking. Conclusion: It is demonstrated here the possibility to store mouse spermatozoa for up to five years in a −80°C freezer with no significant differences compared to the storage in LN2 in terms of fertilizing ability, sperm viability, intracellular calcium concentration, membrane lipid disorder, and mitochondrial activity. [ABSTRACT FROM AUTHOR]

Published
2021
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2. The impact of five years storage/biobanking at -80°C on mouse spermatozoa fertility, physiology, and function

Author

Claudio Ortolani, Mario D'Atri, Marco Marchisio, Renata Paoletti, Stefano Papa, Barbara Barboni, Ferdinando Scavizzi, Marina Ramal-Sanchez, Nicola Bernabò, Sabrina Putti, Paola Lanuti, Marcello Raspa, and Luca Valbonetti

Subjects
Male, Urology, Endocrinology, Diabetes and Metabolism, medicine.medical_treatment, 80°C freezer, flow cytometry, in vitro fertilization, mouse spermatozoa, sperm cryopreservation, Population, Embryonic Development, Fertilization in Vitro, Biology, Calcium in biology, Flow cytometry, Andrology, −80°C freezer, Mice, 03 medical and health sciences, chemistry.chemical_compound, 0302 clinical medicine, Endocrinology, Human fertilization, medicine, Animals, Propidium iodide, Birth Rate, education, Cryopreservation, education.field_of_study, 030219 obstetrics & reproductive medicine, In vitro fertilisation, medicine.diagnostic_test, Embryo, Spermatozoa, Sperm, Mice, Inbred C57BL, Reproductive Medicine, chemistry
Abstract

Background We previously demonstrated how mouse spermatozoa can be efficiently stored for two years in a -80°C freezer, maintaining their ability to fertilize mouse eggs. Objectives The main objective here was to evaluate the effects of five years at -80°C compared to liquid nitrogen storage (LN2 , control condition) on mouse sperm viability, physiological parameters, and fertilization capacity. Materials and methods Three different strains were used: C57BL/6N, C57BL/6J and CD1. Flow cytometry experiments were performed to analyze sperm viability (SYBR-14 + Propidium Iodide +Hoechst33342), the intracellular calcium concentration (Fluo 3-AM), the membrane lipid disorder (Merocyanine 540), and the mitochondrial activity (MitoTracker Red) in live spermatozoa. The in vitro fertilization (IVF) was used to evaluate the sperm fertilizing ability. Results Flow cytometry analysis showed that the percentage of live cells are reduced in B6N and B6J, but not in CD1 mice. However, in the live population no differences in terms of intracellular calcium concentration, membrane lipid disorder, and mitochondrial activity were reported when comparing both biobanking methods. Spermatozoa stored at -80°C for 5 years successfully fertilized the eggs and developed mouse embryo normally both in culture and in vivo, generating live pups with no differences compared to control samples stored in LN2 . Discussion Long-term mouse sperm storage at -80°C (five years) could be considered an ideal alternative to the most common LN2 approach, giving economical and logistic advantages. Moreover, the precise information originated from the flow cytometry analysis stands up this technique as an optimal strategy to evaluate the sperm quality and ranking. Conclusion It is demonstrated here the possibility to store mouse spermatozoa for up to five years in a -80°C freezer with no significant differences compared to the storage in LN2 in terms of fertilizing ability, sperm viability, intracellular calcium concentration, membrane lipid disorder, and mitochondrial activity.

Published
2021

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