Slide Preparation and Staining Procedures for Reliable Results Using Computerized Morphology

Lacquet, Françoise A.; Kruger, Thinus F.; Toit, T.C. du; Lombard, Carl; Sarmiento, C.A. Sánchez; Villiers, Anniza de; Coetzee, Kevin

doi:10.3109/01485019608987089

Cited by 34 publications

(26 citation statements)

References 15 publications

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“…Thin smears facilitated sperm morphology assessment by avoiding sperm cell overlap and ensuring that the sperm were scattered at the same focal depth. The airdried slides were stained with Diff-Quik stain (Merck Diagnostica, Darmstadt, Germany) as previously described (30). Briefly, the slides were fixed for 20 seconds in solution 1, stained for 10 seconds in solution 2 and for 10 seconds in solution 3, and washed in water.…”

Section: Sperm Morphology Assessmentmentioning

confidence: 99%

A novel approach for the selection of human sperm using annexin V-binding and flow cytometry

Hoogendijk

Kruger

Bouic

et al. 2009

Fertility and Sterility

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Section: Sperm Morphology Assessmentmentioning

confidence: 99%

A novel approach for the selection of human sperm using annexin V-binding and flow cytometry

Hoogendijk

Kruger

Bouic

et al. 2009

Fertility and Sterility

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“…Sperm concentrations for CAMA slides were adjusted to the optimal number as previously established in our laboratory (Lacquet et al, 1996). In this phase we compared the two optimal staining methods as identified in phase one.…”

Section: Experimental Designmentioning

confidence: 99%

Effects of different staining and washing procedures on the results of human sperm morphology evaluation by manual and computerised methods

et al. 2009

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The purpose of this study was to investigate the effect of different staining and washing procedures on the results of human sperm morphology evaluation by manual and computerised methods. Furthermore, it was intended to find the staining and washing combination which would provide optimal readability for computer-assisted sperm morphology evaluations. In phase one, four staining methods were evaluated for smears prepared from the resulting samples following a two times washing procedure. In phase two, 20 semen samples were used to compare the DS-Quik and Papanicolaou staining methods, following one and two washes. All manual readings, of Papanicolaou and Diff-Quik stained smears, were comparable with each other, with means between 7.3% and 7.9% normal spermatozoa. All the manual readings were also comparable to the computer readings of the DS-Quik slides following one and two washes with means of 9.0% and 5.9%, respectively. However, due to the higher computer readings found for the Papanicolaou stained smears, with means of 13.9% and 13.5% following one and two washes, respectively, a statistically significantly difference between overall computer and manual readings was found (Wilks' Lamda, P= 0.0002). Taking all data into consideration, it could be concluded that the one wash DZ-Quik stained smears was the optimal preparation method for computerised sperm morphology evaluation, comparing favourably with manual evaluations.Correspondence: R. Menkveld PhD, Andrology Laboratory E3, Tygerberg Hospital, 7505 Tygerberg, South Africa.

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“…When used both with standardized methods and with variables for analysis [21,23,24], these machines provide high repeatability and precision compared with subjective npg morphological evaluation [25,26]. However, problems of cell recognition, image digitization [2], and the effects of staining procedures [11] cannot be avoided and the process is time-consuming.…”

Section: Introductionmentioning

confidence: 99%

Dimensions of human ejaculated spermatozoa in Papanicolaou-stained seminal and swim-up smears obtained from the Integrated Semen Analysis System (ISAS®)

Bellastella¹,

Cooper²,

Battaglia³

et al. 2010

Asian J Androl

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Objective measurements are required for computer-aided sperm morphometric analysis (CASMA) machines to distinguish normal from abnormal sperm heads. The morphometric characteristics of spermatozoa in 72 samples of semen and of spermatozoa from 72 other semen samples after swim-up were quantified by the semi-automated Integrated Sperm Analysis System (ISAS) computer-aided system, which measured the sperm head parameters length (L), width (W), area (A), perimeter (P), acrosomal area (Ac), and the derived values L/W and P/A. For each man a homogeneous population of distributions characterized seminal spermatozoa (7 942 cells: median values L 4.4 µm, W 2.8 µm, A 9.8 µm 2 , P 12.5 µm, Ac 47.5%, L/W 1.57, P/A 1.27), and there was no significant difference in within-and among-individual variation. Different men could have spermatozoa of significantly different dimensions. Head dimensions for swim-up spermatozoa from different men (4 812 cells) were similar to those in semen, differing only by 2%-5%. The values of L, W and L/W fell within the limits given by the World Health Organization (WHO). Although these samples were not biologically matched, linear mixed-effects statistical analyses permitted valid comparison of the groups. A subpopulation of 404 spermatozoa considered to fit the stringent criteria of WHO 'normal' seminal spermatozoa from both semen and swim-up were characterized by median values (and 95% confidence intervals) of L, 4.3 µm (3.8-4.9), W, 2.9 µm (2.6-3.3), A, 10.2 µm 2 (8.5-12.2), P, 12.4 µm (11.3-13.9), Ac, 49% (36-60), L/W, 1.49 (1.32-1.67) and P/A, 1.22 (1.11-1.35). These median values fall within the 95th centile confidence limits given by WHO, but the confidence intervals for L and W were larger. Although these differences in head dimensions among men and after swim-up could be detected by CASMA, the small differences make it unlikely that technicians would be able to distinguish them. The values could be used as default sperm head values for the CASMA machine used here.

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Slide Preparation and Staining Procedures for Reliable Results Using Computerized Morphology

Cited by 34 publications

References 15 publications

A novel approach for the selection of human sperm using annexin V-binding and flow cytometry

A novel approach for the selection of human sperm using annexin V-binding and flow cytometry

Effects of different staining and washing procedures on the results of human sperm morphology evaluation by manual and computerised methods

Dimensions of human ejaculated spermatozoa in Papanicolaou-stained seminal and swim-up smears obtained from the Integrated Semen Analysis System (ISAS®)

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