A comparison of motility and head morphology of sperm using different semen processing methods and three different staining techniques

[摘要] ENGLISH ABSTRACT: Sperm morphology remains an important parameter in the prediction of fertility, bothin vivo and in vitro. However, there remains a considerable level of concernsurrounding the true potential of this parameter due to the lack of standardization ofdifferential staining techniques used for the evaluation of sperm morphology. Thisstudy aimed at investigating two commonly used staining techniques, Rapidiff® (RD)and Papanicolaou (PAP), along with a new commercially available stain, SpermBlue®(SB), in the evaluation of sperm morphometry and morphology. Results indicated thatsignificant differences in sperm morphometry exist due to the use of the stainingtechniques. Findings further indicated that RD causes sperm head swelling while PAPcauses sperm head shrinkage. Results obtained using the SB staining technique haveindicated measurements closest to that which would be obtained through theevaluation of fresh, unstained sperm. The lack of standardization and the differenteffects various stains have on sperm structure and overall sperm morphologyevaluation should raise a level of concern, particularly when evaluating patients withborderline morphology. Based on this, the use of the SB staining technique isrecommended over RD and PAP for effective and accurate morphology evaluation. Infurther support of this technique, SB was shown to be quick and simple in method,and allowed for the easy detection of sperm by computer aided sperm analysis(CASA) systems such as the Sperm Class Analyzer (SCA®).The second aim of this study was to examine the concentration, morphology andmotility of the resultant sperm populations following semen preparation using thePureSperm® density gradient and swim-up techniques. Semen preparation is anessential step in any fertility treatment protocol, and it is important that the spermobtained following semen preparation has sperm morphology and motilitycharacteristics capable of improving assisted fertility success rates. Currently, thePureSperm® density gradient and sperm swim-up are the most widely employedtechniques in fertility clinics. Although there is sufficient evidence to suggest they areeach effective at extracting sperm with improved quality from neat semen, thereremains insufficient evidence to suggest which of these two techniques is superior.The present investigation revealed that both sperm preparation methods were effective at improving sperm morphology and motility, however to varying degrees. The swimupmethod yielded a population of sperm with superior motility and morphologywhen assessed according to World Health Organisation (WHO) criteria, while thePureSperm® density gradient technique isolated a higher percentage of normal sperm,according to both WHO and Tygerberg strict criteria, with motility better than that ofneat semen. Although results obtained via the swim-up method suggest it would bebest for use in in vitro fertilization (IVF), the very low concentration of sperm isolatedvia this method remains a significant draw-back. The PureSperm® density gradientseparation technique on the other hand is capable of isolating larger quantities ofsperm, which is likely to be of more benefit with fertility treatments requiring largerquantities of sperm. Based on these findings, the use of PureSperm® density gradienttechnique is recommended, due to its ability to isolate large quantities of good qualitysperm. However, a swim-up may still be of use when performing fertility treatmentusing a sperm sample which possesses a high concentration and motility.

[发布机构] Stellenbosch University