Dear Hetal,

I have read two publications one in 2013 and the other in 2015 that studied the effect of semen storage in liquid nitrogen and both have concluded that up to one year there is no DNA fragmentation or membrane integrity change:

1-Asian Pacific Journal of Reproduction 2015; 4(1): 22-25

Effects of long term storage of semen in liquid nitrogen on the viability,

motility and abnormality of frozen thawed Frisian Holstein bull

spermatozoa

Abdul MALIK1*

, Muhammad LAILY2

, Muhammad Irwan ZAKIR

Objective: To evaluate effects of long term storage of semen in liquid nitrogen on the motility, concentration, viability, and abnormality of frozen-thawed. Methods: A total of four Friesian Holstein bulls were used for this study. One hundred forty semen straws with produced during period from 2008 to 2013 and stored in the liquid nitrogen at the AI center were used in the research. The sample straw was divided into six groups; each group consist 20 semen straws. For group one all straw semen was produced on the 2013 with storage in liquid nitrogen as long as

one year, the group 2, 3, 4, 5, and 6 were produced on the 2012, 2011, 2010, 2009 and 2008 with storage in liquid nitrogen as long as 2, 3, 4, 5, and 6 year, respectively.

Results: The viability of thawed sperm was not significantly different decreased (P>0.05) between storage on the 1 year and storage on the 2 years. Whereas, the viability was significantly different (P0.05) on the storage 1, 2 and 3 years. Whereas, the motility was significantly different (P0.05) on the storage 1, 2 and 3 years. Whereas, the abnormality was significantly increased (P0.05) during storage in liquid nitrogen as long as six years.

Conclusions: Based on the results in these experiments,

it may be concluded that concentration sperm during one year storage in liquid nitrogen resulted in similar concentration storage as long as six years. However, the viability and motility sperm thawed storage in liquid nitrogen during six years was lower than storage on the 1 and 2 years.

For more details, see attached file.

2-Effects of Storage Duration on the Quality and DNA Integrity of Nili-Ravi Bull Spermatozoa

Frozen and Stored in Liquid Nitrogen

Muhammad Shoaib, Nazir Ahmad*, Maqbool Ahmad, Ijaz Ahmad, Sajid Iqbal1

and Shamim Akhter2

In the present study, effects of storage duration on the quality and DNA integrity of

Nili-Ravi buffalo bull spermatozoa frozen and stored in liquid nitrogen for up to

three years were investigated. According to duration of storage, the semen samples

were categorized into 10 groups, viz. i) 0 day, ii) one day, iii) two weeks, iv) one

month, v) three months, vi) six months, vii) nine months, viii) one year, ix) two

years and x) three years. A total of 50 straws, with 5 straws for each of the 10

groups were used. After thawing for 30 seconds at 37°C, these samples were

evaluated for sperm motility, livability, livability index, plasma membrane integrity,

acrosome damage and DNA damage. At 0 day, mean values for sperm motility (%),

livability (hrs), livability index, plasma membrane integrity (%), acrosome damage

(%) and DNA damage (%) were 53±1.22, 6.0±0.0, 130±5.2, 45.4±1.6, 3.8±0.6 and

2.2±0.4. After 3 years of storage, the corresponding values were 39±3.32, 4.4±0.2,

78.5±5.7, 28.2±2.2, 12.0±0.7 and 5.8±0.6. Sperm motility and livability remained

unaffected up to 3 months of storage, followed by a decrease at 6th month, while

absolute index of livability and membrane integrity started decreasing at 1 month

(P

Thanks Rafik.