A fertility-oriented method for histological processing of testicular biopsies in men with azoospermia.

The present study was designed to evaluate whether tissue preparation by glutaraldehyde and glycol methacrylate (G/GMA) improves the diagnostic assessment of testicular biopsies from azoospermic men when compared to the standard tissue preparation using Bouin's solution and paraffin. We prospectively included a total of 21 testicular biopsies of sexually mature men aged 29-50 years with infertility and azoospermia. One testicular biopsy fragment from each patient was processed by the G/GMA method, whereas another tissue fragment was contemporarily processed by the conventional Bouin/paraffin (B/P) method. The G/GMA method provided better resolution of cytological details of the seminiferous epithelium, changing the final diagnosis in four cases. The medians of Bergmann's spermatogenesis scores were 0.25 (interquartile range 0.04-0.88) for B/P preparations and 0.79 (interquartile range 0.17-0.96) for G/GMA preparations. Both techniques allowed accurate prediction of sperm recovery from the biopsies (B/P, area under the receiver operating characteristics [ROC] curve 0.88, 95% confidence interval [CI] 0.75-1.00; G/GMA, area under the ROC curve 0.94, 95% CI 0.86-1.00). We conclude that human testicular biopsy preparation with G/GMA improved image resolution under light microscopy and produced more reliable results for the evaluation of spermatogenesis in comparison with B/P, allowing a more precise fertility-oriented diagnosis in azoospermic men.Abbreviations: B/P: Bouin/paraffin; GMA: glycol methacrylate; G/GMA: glutaraldehyde and glycol methacrylate; ICSI: intracytoplasmic sperm injection; OA: obstructive azoospermia; NOA: nonobstructive azoospermia; TESE: testicular sperm extraction.

Influence of three different histological methods on the morphology and morphometrical data in human testis.

Coagulant fixatives and paraffin embedding were widely used in the past for histomorphometrical evaluations of the human testis under physiological and pathological conditions. However, new methods are applied nowadays using better combinations of fixatives and plastic resins as embedding media, improving cell and tissue structural preservation. In an attempt to compare old and new data, the present study evaluated histomorphometrical data obtained from human testis after three different histological processing methods: Bouin/paraplast, glutaraldehyde/glycol methacrylate and glutaraldehyde/araldite. The morphometrical parameters were not affected by glutaraldehyde fixation after both resin embedding (methacrylate or araldite). On the other hand, Bouin/paraplast embedding lead to tissue shrinkage, which could give rise to misinterpretations on the measurements performed. Since some germ and somatic cells recognition do not depend upon high resolution techniques, counting of such cell types could be performed even using routine Bouin/paraplast protocols. Thus, the morphometrical analyses relying on cell recognition were not affected by the methods here applied, however, when metric measurements were applied, the obtained results could not be promptly compared. However, if the study requires confident spermatogonial identification for kinetics evaluation, glutaraldehyde/araldite processing is highly recommended.