Human Sperm Cells After Purification Using SCLB Can Be Stored at 4o, -20o, or -80oC Before Small RNA Isolation
DOI:
https://doi.org/10.24293/ijcpml.v26i2.1530Keywords:
Sperm RNA analysis, purified sperm cells, storage temperatureAbstract
There have been many studies about pre-analysis for sperm RNA examination which compared sperm purification methods, RNA isolation methods, sequencing methods, and semen storage before analysis. However, there is a lack of studies that determine the ideal storage temperature after sperm cell purification before RNA analysis, especially small RNA analysis. The aim of this study was to determine the preferred storage temperature for human sperm cells after sperm purification using Somatic Cell Lysis Buffer (SCLB) before sperm small ribonucleic acid (RNA) isolation and analysis. This
study was a true laboratory experiment using the post-test only control group design. The samples were 13 fresh human semen that has been purified using SCLB. The sperm cells were then diluted and divided into four aliquots with different treatments. The first aliquot that served as a control group was immediately purified while the other three aliquots were 0 0 0 stored for seven days at different temperatures as follows: 4 C, -20 , and -80 C. After the small RNA isolation, RNA level between each group was compared. Micro volume spectrophotometer measured RNA level. The median of small RNA
6 yields of the control group was 49.8 (5.33-522.46) ng/10 sperm cells. There was no significant difference in median of small RNA yields of the control group and that of other groups. The median of the other groups with storage temperature 0 0 0 6 of 4 C, -20 , and -80 C was 41.09 (7.03-1448.31), 65.95 (7.99-301.16), and 76.42 (10.45-434.25) ng/10 sperm cells, respectively (p-value= 0.314; α=5%). This condition suggested that after purification using SCLB, human sperm cells can be 0 0 0 stored at temperatures of 4 C, -20 , or -80 C for seven days, depending on each laboratory facility.
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References
Agarwal A, Mulgund A, Hamada A, Chyatte MR. A unique view on male infertility around the globe. Reprod Biol Endocrinol. 2015;13(37):1–9.
Anton E, Krawetz SA. Spermatozoa as biomarkers for the assessment of human male infertility and genotoxicity. Syst Biol Reprod Med. 2012;58:41–50.
Ramasamy R, Scovell JM, Kovac JR, Cook PJ, Lamb DJ, Lipshultz LI. Fluorescence in situ hybridization detects increased sperm aneuploidy in men with recurrent pregnancy loss. Fertil Steril. 2015;103:906–10.
Agarwal A, Majzoub A, Esteves SC, Ko E, Ramasamy R, Zini A. Clinical utility of sperm DNA fragmentation testing: practice recommendations based on clinical scenarios. Transl Androl Urol. 2016;5(6):935–50.
Jodar M, Sendler E, Moskovtsev SI, Librach CL, Goodrich R, Swanson S, et al. Absence of sperm RNA elements correlates with idiopathic male infertility. Sci Transl Med. 2015;7(295):1–8.
Ellervik C, Vaught J. Preanalytical Variables Affecting the Integrity of Human Biospecimens in Biobanking. Clin Chem. 2015;61(7):914–34.
Mao S, Goodrich RJ, Hauser R, Schrader SM, Chen Z, Krawetz SA. Evaluation of the effectiveness of semen storage and sperm purification methods for spermatozoa transcript profiling. Syst Biol Reprod Med. 2013;59:287–95.
Barragan M, Martınez A, Llonch S, Pujol A, Vernaeve V, Vassena R. Effect of ribonucleic acid (RNA) isolation methods on putative reference genes messenger RNA abundance in human spermatozoa. Andrology. 2015;3:797–804.
Mao S, Sendler E, Goodrich RJ, Hauser R, Krawetz SA. A comparison of sperm RNA-seq methods. Syst Biol Reprod Med. 2014;60(5):308–15.
WHO. WHO laboratory manual for the Examination and processing of human semen. 5 th. Cooper TG, editor. Geneva: WHO; 2010.
Hubel A, Spindler R, Skubitz APN. Storage of human biospecimens: Selection of the optimal storage temperature. Biopreserv Biobank. 2014;12(3):165–75.
Ikeda K, Ichihara K, Hashiguchi T, Hidaka Y, Kang D, Maekawa M, et al. Evaluation of the short-term stability of specimens for clinical laboratory testing. Biopreserv Biobank. 2015;13(2):135–43.
Olivieri EHR, Franco L de A, Pereira RG, Mota LDC, Campos A nio HJFM, Carraro DM. Biobanking practice: RNA storage at low concentration affects integrity. Biopreserv Biobank. 2014;12(1):46–51.
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