Evaluation of black glutinous rice (Oryza sativa L) extract as a novel nuclear stain for human sperm head assessment by microscopic examination / 대한생식의학회지

OBJECTIVE: To compare black rice (Oryza sativa L) extract with three different staining methods for human sperm head assessment. METHODS: Semen samples were collected from 34 volunteers. Four smears of each ejaculate were prepared for staining using the rapid Papanicolaou (PAP) stain, SpermBlue, DipQuick, and black rice extract. The percentage of defective sperm heads (mean±standard deviation) was compared. RESULTS: Black glutinous rice extract, a natural dye, was used instead of hematoxylin to stain the nuclei of the sperm heads. The percentage of defective sperm heads showed a significant difference between black rice extract and DipQuick (p=0.000). In contrast, black rice extract and rapid PAP showed no statistically significant difference (p=0.974). A strong correlation (r =0.761) was found between the findings obtained using rapid PAP and black rice extract. In contrast, a weak correlation (r =0.248) was obtained between DipQuick and black rice extract for the percentage of defective sperm heads. CONCLUSION: The results showed good agreement and a strong correlation between the rapid PAP and black rice extract stains. The advantages of black rice extract as a novel substitute for hematoxylin for nuclear staining include ease of preparation, local availability, and favorable nuclear staining properties. Further studies could also focus on comparing staining techniques in clinical samples.

Comparative study of semen quality between pre-washed and post-washed with 3 sperm preparation media

The semen preparation medium that yields good semen quality is one among various important factors in the process of infertility treatment. The objective of this study was to compare the semen analysis parameters before and after preparation with 3 semen preparation media i.e. PureSperm® (Nidacon, Gothenburg, Sweden), Sil-Select Plus™ (Fertipro, Beernem, Belgium) and SpermGrad™ (Vitrolife, Gothenburg, Sweden) for enhancement of good semen quality.The total of 28 males of infertile couples attending Fertility Clinic of Thammasat University Hospital during the year 2007 were studied. Their semen (pre-washed) were analyzed for semen analysis parameters according to WHO¹ and Kruger’ strict criteria² and then equally divided into 3 aliquots and processed with 3 semen preparation media followed by semen analysis once again (post-washed). The sperm concentration, motility, progressive motile concentration (PMC), morphology, DNA damage and protamine deficiency of before and after preparation were compared by paired t-test with significance at P \< 0.05.The mean percentage of sperm motility, PMC, rapid motility and normal morphology were significantly increased where the mean percentage of slow motility, sperm head defect, sperm concentration and protamine deficiency were significantly decreased after preparation with all 3 media. The DNA damaged sperm was significantly decreased in the post-washed with PureSperm® and Sil-Select Plus™ but significantly increased with SpermGrad™.From the results obtained in this study, it could be concluded that PureSperm® may be suitable for both IUI and IVF techniques, since it yielded the best motile sperm and it may be also good for ICSI because it gave low percentage of DNA damaged sperm. Sil-Select Plus™ is probably best for ICSI since it gave lowest percentage of DNA damage sperm, whereas SpermGrad™ had highest percentage of DNA damage, thus, it is surely not suitable for ICSI procedure. Anyhow, SpermGrad™ may be moderately good for IUI since it had high percentage of motility and of normal morphology with lowest percentage of head defect but percentage of abnormal embryo may be increased.