Theta and alpha oscillatory signatures of auditory sensory and cognitive loads during complex listening.

The neuronal signatures of sensory and cognitive load provide access to brain activities related to complex listening situations. Sensory and cognitive loads are typically reflected in measures like response time (RT) and event-related potentials (ERPs) components. It's, however, strenuous to distinguish the underlying brain processes solely from these measures. In this study, along with RT- and ERP-analysis, we performed time-frequency analysis and source localization of oscillatory activity in participants performing two different auditory tasks with varying degrees of complexity and related them to sensory and cognitive load. We studied neuronal oscillatory activity in both periods before the behavioral response (pre-response) and after it (post-response). Robust oscillatory activities were found in both periods and were differentially affected by sensory and cognitive load. Oscillatory activity under sensory load was characterized by decrease in pre-response (early) theta activity and increased alpha activity. Oscillatory activity under cognitive load was characterized by increased theta activity, mainly in post-response (late) time. Furthermore, source localization revealed specific brain regions responsible for processing these loads, such as temporal and frontal lobe, cingulate cortex and precuneus. The results provide evidence that in complex listening situations, the brain processes sensory and cognitive loads differently. These neural processes have specific oscillatory signatures and are long lasting, extending beyond the behavioral response.

A cortical biomarker of audibility and processing efficacy in children with single-sided deafness using a cochlear implant.

The goals of the current study were to evaluate audibility and cortical speech processing, and to provide insight into binaural processing in children with single-sided deafness (CHwSSD) using a cochlear implant (CI). The P1 potential to acoustically-presented speech stimuli (/m/, /g/, /t/) was recorded during monaural [Normal hearing (NH), CI], and bilateral (BIL, NH + CI) listening conditions within a clinical setting in 22 CHwSSD (mean age at CI/testing 4.7, 5.7 years). Robust P1 potentials were elicited in all children in the NH and BIL conditions. In the CI condition: (1) P1 prevalence was reduced yet was elicited in all but one child to at least one stimulus; (2) P1 latency was prolonged and amplitude was reduced, consequently leading to absence of binaural processing manifestations; (3) Correlation between P1 latency and age at CI/testing was weak and not significant; (4) P1 prevalence for /m/ was reduced and associated with CI manufacturer and duration of CI use. Results indicate that recording CAEPs to speech stimuli in clinical settings is feasible and valuable for the management of CHwSSD. While CAEPs provided evidence for effective audibility, a substantial mismatch in timing and synchrony of early-stage cortical processing between the CI and NH ear remains a barrier for the development of binaural interaction components.

Spermiogenesis defects in human: detection of transition proteins in semen from some infertile men.

Semen samples from 60 infertile men were examined by flow cytometry following propidium iodide staining. Of these, 23 samples contained young haploid cells. Transition proteins (TP1 and/or TP2) were detected in 12 of these, using immunohistochemical staining. The presence of TPs in spermatids in semen indicates inhibition in the differentiation pathway from round spermatids to spermatozoa. Cells of this type were found in semen from patients with nonobstructive azoospermia, severe to extreme cases of oligozoospermia, asthenozoospermia and teratozoospermia.

Flow cytometry of human semen: a preliminary study of a non-invasive method for the detection of spermatogenetic defects.

BACKGROUND: The pathway of spermatogenesis involves the conversion of diploid stem cells (spermatogonia) to tetraploid primary spermatocytes, followed by meiosis and two cell divisions, first forming diploid secondary spermatocytes and then haploid round spermatids. Differentiation of round spermatids results in spermatozoa containing condensed chromatin. It has long been known that semen from patients with non-obstructive azoospermia or oligospermia contains small numbers of immature germinal cells. In this article, a flow cytometric procedure is described for assessing defects in spermatogenesis by identifying the ploidy of those immature cells. METHODS: Cells in semen samples from 44 infertile patients and 14 controls were stained with propidium iodide, which displays red fluorescence when intercalated between bases in double-stranded DNA. The resulting cell suspension was examined by quantitative flow cytometry, with excitation by laser light (488 nm) and red fluorescence recorded on a logarithmic scale to allow easy differentiation between intensities of tetraploid, diploid and haploid round spermatids, and spermatozoa containing condensed chromatin. RESULTS: The flow cytometric method differentiated between cases of 'Sertoli cell-only' syndrome (complete absence of tetraploid and haploid cells) and cases where spermatogenesis was blocked in meiosis or in spermiogenesis. Flow cytometric histograms from semen samples from normozoospermic, oligozoospermic and azoospermic patients fell into patterns that correlated well with the results obtained from testis histology findings. CONCLUSIONS: The method described may serve as a simple, non-invasive and reliable assay to help clinicians counsel patients with severe male infertility before referring them for testicular surgery to locate spermatozoa for ICSI.

Membrane fouling and selectivity mechanisms in effluent ultrafiltration coupled with flocculation.

Membrane filtration is adequate for producing disinfected clear water suitable for various kinds of applications. However, fouling of membranes is the main limitation. The scope of the present study is to examine the effect of iron coagulation of primary wastewater effluent on membrane filtration, in parallel to fouling characterization of the iron itself. The fouling of ultrafiltration membranes by colloidal iron hydroxide-oxide has been studied by measuring the pore streaming potential of PES UF membrane. pH 5.5 (charge neutralization zone) provided better removal and lower fouling intensity than pH 7.8 (sweep coagulation zone), but the internal clogging at acidic pH was higher. Fouling of the membrane as measured by flux reduction was usually accompanied by a positive change in zeta potential and iso-electric point (IEP) of the membrane. An initially large change in zeta potential (without charge reversal) was seen even after relatively small amounts of iron solution were filtered through the membrane. A control experiment showed this is not due to iron adsorption equilibrium, but should probably be attributed to fouling. Change in zeta potential, can be used as an indicator for commencement of fouling even for small flux reductions. UF membrane critical flux after iron filtration can be evaluated more accurately by zeta potential than pressure drop or change in iron concentration.

[Evaluation of sperm samples from infertile men by flow cytometry].

INTRODUCTION: Human spermatogenesis begins at adolescence and continues throughout life. This process includes morphologic, cytologic and biological changes, leading to the formation of mature spermatozoa. Male infertility may be caused by several reasons, including oligozoospermia at variable degrees and complete absence of mature spermatozoa. Routine spermatogram, measuring sperm counts, motility and morphology, might not provide complete information in the evaluation of these cases. This study is aimed to evaluate the possible use of flow cytometry in the identification of different sperm cell populations in sperm samples obtained from infertile men, and in determining the different cell types in various groups of infertile men. MATERIALS AND METHODS: Sperm samples from normal and infertile men (the latter were azoospermic or oligoteratozoospermic OTA) underwent flow cytometry analysis, after preparation with TNE buffer and staining with Propidium Iodide. The separation of germinal cells into different populations, according to their DNA content and chromatin condensation, was evaluated. The WINMDI (http://fac.-scripps.edu, J. Trotter) software was used for data analysis. RESULTS: Flow cytometric analysis enabled identification of several cell populations in sperm samples, including haploid, diploid and tetraploid cells. Certain cellular distribution patterns were observed in sperm samples from infertile men: mature haploid cells, diploid cells, domination of tetraploid or non-mature haploid cells, and combination of these patterns. These patterns appeared in a statistically different manner among fertile and infertile men; the median value of mature haploid cells was higher in normal men (91%, compared to 85% in the OTA group and 0% in the azoospermic men), while the median value of diploid and tetraploid cells was higher in azoospermic men (72% and 8.5% respectively, compared to only 1% and 0% in normal men). CONCLUSIONS: These findings suggest that flow cytometry of sperm samples may serve as a non-invasive tool for investigations of male infertility and for identification of appropriate candidates for interventional treatment.

Factors influencing the outcome of ICSI in patients with obstructive and non-obstructive azoospermia: a comparative study.

BACKGROUND: Factors influencing success of sperm retrieval in azoospermic patients and outcome of ICSI were evaluated. METHODS AND RESULTS: Uni- and multifactorial analysis were performed using logistic and stepwise analysis, following surgical sperm retrieval by percutaneous epididymal sperm aspiration (55 cycles) or testicular sperm extraction (142 cycles) in 52 and 123 patients with obstructive azoospermia (OA) and non-obstructive azoospermia (NOA) respectively. ICSI cycles using fresh or cryopreserved-thawed sperm were included. Sperm were retrieved to allow ICSI in 100 and 41% of OA and NOA patients, with no significant correlation with patients' age or FSH level. Occurrence of pregnancy was significantly correlated with female age (90th quantile: 38 years), number of oocytes retrieved (10th quantile: five oocytes) and number of oocytes injected (10th quantile: four oocytes). Sperm origin (epididymal versus testicular), status (fresh or thawed), male partner's age, and serum FSH had no significant effect upon implantation rate, pregnancy rate per embryo transfer or spontaneous miscarriage rate. CONCLUSIONS: In OA patients ICSI should be planned in conjunction with surgical sperm retrieval. In contrast, the lack of efficient non-invasive parameters to predict sperm retrieval in NOA suggests that elective surgical sperm retrieval may be offered to these patients prior to ovarian stimulation of their partners, especially when donor back-up is not an alternative. Female factors such as age and ovarian reserve have significant impact upon clinical success rates.

Influence of a short or long abstinence period on semen parameters in the ejaculate of patients with nonobstructive azoospermia.

OBJECTIVE: To compare the effect of a short (4 days) or a long (14 days) abstinence period on sperm retrieval by extended sperm preparation in patients with nonobstructive azoospermia scheduled for testicular biopsy and intracytoplasmic sperm injection (ICSI). DESIGN: A prospective case control study. SETTING: Male infertility clinic in a university hospital. PATIENT(S): Fifty male patients with nonobstructive azoospermia, scheduled for testicular biopsy for ICSI. INTERVENTION(S): Diagnosis of nonobstructive azoospermia and a thorough microscopic search for sperm cells (extended sperm preparation). MAIN OUTCOME MEASURE(S): The number of sperm cells collected, sperm motility, and total motile sperm count after short and long abstinence periods. RESULT(S): There was a significant difference between long and short abstinence with an increase in sperm count (log-to-log transformed analysis of variance P<.025) and total motile sperm (P<.025 analysis of variance, P<.02 paired Student's t-test) in the former group, but no significant change in sperm motility (Wilcoxon and paired Student's t-test). In 18 patients, sperm concentration and sperm motility were similar in a second collection, done after the same abstinence period, compared with the same parameters in the first sample. When at least 10 motile sperm were defined as the cutoff number, allowing ICSI without testicular biopsy, no significant differences were found between the two abstinence periods. No clinical or laboratory male characteristic could predict the detection of 10 motile sperm by extended sperm preparation either after a short or a long abstinence period. CONCLUSION(S): Sperm count and total motile sperm were increased after a long abstinence period, with no change in sperm motility. No additional advantages were conferred by long abstinence as opposed to short abstinence when 10 motile sperm were defined as the cutoff number for ICSI. The recommended period of abstinence for extended sperm preparation and ICSI, whether short or long, should be individualized for each patient.

In vitro susceptibility of recent clinical isolates of Chlamydia trachomatis to macrolides and tetracyclines.

We tested the in vitro activity of clarithromycin, azithromycin, roxithromycin, erythromycin, doxycycline, and tetracycline against 50 clinical isolates of Chlamydia trachomatis. The minimal inhibitory concentrations (MICs) and the minimal bactericidal concentrations (MBCs) were determined in a tissue culture system using cycloheximide treated McCoy cells. MIC values for all the isolates were < or =0.015 microg/ml for clarithromycin, < or =0.125 microg/ml for roxithromycin and azithromycin, and < or =0.25 microg/ml for erythromycin and doxycycline. Almost half of the isolates (44%) were inhibited only by a concentration of 0.5 microg/ml of tetracycline. MBC as high as 4 microg/ml was displayed by doxycycline and tetracycline against 8% and 4% of the isolates respectively of the agents recommended by the Center for Disease Control as drugs of choice for the treatment of chlamydial infections, azithromycin exhibited a markedly better in-vitro activity than did erythromycin and doxycycline.

Outcome of surgical treatment for superficial dyspareunia from vulvar vestibulitis.

OBJECTIVE: To evaluate the results of surgical treatment for superficial dyspareunia, as manifested by patient satisfaction, as well as epidemiologic characteristics of women with this medical problem. STUDY DESIGN: A questionnaire was sent to 69 women six months after the operation. It included questions about treatment before surgery and the impact of pain on the sexual relationship before and after the operation. Demographic, social and general health data were recorded before the operation. All patients returning the questionnaire were examined. RESULTS: Fifty-four (78%) patients replied. Half of those abstained from sexual relations before surgical treatment. Sixty-seven percent of patients required more than six visits to various physicians, before vestibulitis was diagnosed. Prior to surgery, 80% of patients received conservative treatment, whereas after surgery only 34% required it. A moderate to excellent improvement was reported after surgery by 45 (83%) patients. Repeat surgery (n = 7) resulted in further improvement in four patients. There were no major operative complications. Forty-five patients (83%) were satisfied with the results and would recommend the surgery to other women with this clinical problem. CONCLUSION: Surgical treatment for superficial dyspareunia from vestibulitis is quite safe and results in a high rate of patient satisfaction.

Sperm ultramorphology as a pathophysiological indicator of spermatogenesis in males suffering from varicocele.

Varicocele of spermatic veins is considered to be one of the major causes of male infertility associated with reduction of sperm quality. The pathophysiology of this condition is not yet completely understood. The aim of this study was to shed light on the pathophysiology of varicocele by identifying semen parameters, especially sperm ultramorphology, which improve following high ligation of the spermatic vein. Seventy-five males with diagnosed varicocele were included in this study. Semen parameters were assessed prospectively using light microscopy, semen biochemistry and sperm quantitative ultramorphological analysis, before high ligation and 3-9 months after high ligation. The control group consisted of twenty-five untreated varicocele patients who underwent two semen examinations within 3-9 months. No statistical difference in any of the examined variables was found between the two examinations in the control group. The treated patients exhibited a significant improvement in sperm density, progressive motility, percentage of normally formed spermatozoa, agenesis of sperm acrosome, chromatin condensation and incidence of amorphous heads compared with the pretreatment condition (P < or = 0.01). In contradiction, no significant improvement was observed following treatment in any of the sperm tail subcellular organelles. It is concluded that varicocele may cause deleterious alterations in early spermatid head differentiation during spermiogenesis and that varicocele patients with a high incidence of sperm acrosome and nucleus malformations are appropriate candidates for varicocele correction.

ART success and in vivo sperm cell selection depend on the ultramorphological status of spermatozoa.

Management of male infertility has recently shifted from treatment of the subfertile man towards techniques of assisted reproduction (ART). This study aimed to evaluate the possible role of the ultramorphological status of the spermatozoon with respect to sperm selection in vivo and prediction of ART success. Ultramorphological sperm parameters were assessed retrospectively for 92 males with sufficient sperm density (10(7) spermatozoa ejaculate-1) whose wives conceived following a stepwise discarding of the female genital tract barriers, using intra-uterine insemination (IUI) (n = 26), in vitro fertilization (IVF) (n = 45) or intracytoplasmic sperm injection (ICSI) (n = 21). In parallel, sperm samples of 71 fertile males were examined. Normal ultramorphology of all head and tail subcellular organelles was found to be essential for the ability of spermatozoa to pass the lower female genital tract. The ultramorphological migration threshold for this barrier is apparently higher than that essential for oocyte fertilization. No specific indication associated with passage through the upper genital tract was found. A high prevalence of axonema defects was found to impair the ability of sperm cells to penetrate the oocyte investment. The natural fertility index, based on routine sperm parameters and the ultrastructural status of the spermatozoon's subcellular organelles was confirmed to be beneficial for directing patients to ART. A discriminative score based on axonema integrity was found to contribute additional information for the first choice decision between conventional ART and ICSI (75% prediction ability). Thus it may be helpful in finding the simplest and least expensive procedure with the greatest long-term chance for pregnancy.

The outcome of intracytoplasmic injection of fresh and cryopreserved epididymal spermatozoa from patients with obstructive azoospermia--a comparative study.

The aim of our study was to compare the outcome of intracytoplasmic sperm injection (ICSI) with fresh and frozen-thawed epididymal spermatozoa retrieved by percutaneous epididymal sperm aspiration (PESA) or microepididymal sperm aspiration (MESA) from patients with obstructive azoospermia. A retrospective analysis of consecutive ICSI cycles was performed, comparing the outcome in 24 patients with obstructive azoospermia undergoing surgical sperm aspiration by MESA (7 cycles) or PESA (17 cycles). In 23 of 24 patients, excess spermatozoa were cryopreserved. Following thawing, 21 ICSI cycles were performed (11 cycles after MESA, 10 after PESA). No statistically significant differences were noted in all parameters examined in ICSI cycles with fresh or cryopreserved spermatozoa from the same patients. Comparing all ICSI cycles with fresh and frozen-thawed epididymal spermatozoa, the rates of two-pronuclear fertilization (56% versus 53%), embryo cleavage (90% versus 86%), implantation (10% versus 14%), clinical pregnancy per embryo transfer (32% versus 37%) and delivery/ongoing pregnancy rate (27% versus 26%) were not statistically different. The cumulative ongoing pregnancy rate per sperm retrieval procedure was 46%, respectively. We conclude that the clinical outcome of ICSI with fresh and frozen-thawed spermatozoa after retrieval by PESA was similar to that by MESA. Epididymal sperm cryopreservation in patients with obstructive azoospermia is feasible and efficient using a simple freezing protocol and should be offered to optimize the yield of pregnancies achieved following such procedures.

Pregnancy and birth after transfer of embryos that developed from single-nucleated zygotes obtained by injection of round spermatids into oocytes.

OBJECTIVE: To use injection of spermatids into oocytes as a mode of infertility treatment in cases in which spermatozoa are not available. DESIGN: Prospective clinical evaluation and case report. SETTING: In Vitro Fertilization Unit, Herzliya Medical Centers, Herzliya-on-Sea, Israel. PATIENT(S): Thirteen couples with male factor infertility in which the male partner lacked spermatozoa in the ejaculate or testicular biopsy samples. INTERVENTION(S): Round spermatid injection and elongated spermatid injection into oocytes. MAIN OUTCOME MEASURE(S): Evaluation of the rate of two-pronucleated and single-nucleated zygote development. RESULT(S): The rate of two-pronucleated zygote development after round spermatid injection and elongated spermatid injection was relatively low (27% and 36%, respectively). Single-nucleated zygotes develop more frequently after round spermatid injection and elongated spermatid injection (35% and 17%, respectively) than after intracytoplasmic sperm injection with mature spermatozoa. A normal pregnancy and childbirth resulted from the transfer of 4 cleaving embryos, each of which developed from a single-nucleated zygote in a round spermatid injection treatment cycle with ejaculated spermatids. CONCLUSION(S): Embryos derived from single-nucleated zygotes after spermatid conception can be viable and give rise to an ongoing clinical pregnancy and childbirth.

In-vitro human spermatozoa nuclear decondensation assessed by flow cytometry.

The process of sperm chromatin decondensation occurs when a spermatozoon enters an ovum. Protamine disulphide bonds are reduced to SH and the polycationic protamines combine with the polyanionic egg protein, nucleoplasmin, thus being stripped from DNA which then combines with histones. Defective chromatin decondensation will thus prevent further development of the male pronucleus. In this study human sperm samples were incubated in vitro at 28 degrees C (using a medium in which the polyanion, heparin, substitutes for nucleoplasmin and beta-mercaptoethanol for egg glutathione) for 10, 20 and 30 min before stopping the reaction with formalin (to 3.6%). The DNA of the fixed cells was stained with Acridine Orange by a one-step method and subjected to flow cytometry and data analysis, in which a zone characteristic of condensed chromatin is outlined on red-green fluorescence contour plots. After 20 min of incubation 97% of the control spermatozoa that were in the mature window (WIN M) had decondensed and moved out of this region. Defects in sperm decondensation were seen in four semen samples of the 20 that were tested. In cases where spermatozoa fail to produce a fertilized egg the cause may lie with defective chromatin quality, including failure of the sperm chromatin to decondense. The method described here is a simple procedure for detecting sperm samples containing such defective cells.

Intracytoplasmic injection of fresh and cryopreserved testicular spermatozoa in patients with nonobstructive azoospermia--a comparative study.

OBJECTIVE: To compare the outcome of intracytoplasmic sperm injection (ICSI) with fresh and frozen-thawed testicular spermatozoa in patients with nonobstructive azoospermia. DESIGN: Retrospective analysis of consecutive ICSI cycles. SETTING: In Vitro Fertilization Unit, Assaf Harofeh Medical Center. PATIENT(S): Eighteen with nonobstructive azoospermia in whom testicular sperm was found after testicular sperm extraction. INTERVENTION(S): Testicular sperm retrieval, cryopreservation, and ICSI with fresh or frozen-thawed testicular spermatozoa. MAIN OUTCOME MEASURE(S): Two-pronuclear fertilization; embryo cleavage rates, mean number of embryos transferred per cycle, and their relative quality, embryo implantation, clinical pregnancy, and ongoing pregnancy rates (PRs) per ET. RESULT(S): No statistically significant differences were noted in all parameters examined between ICSI cycles with fresh or cryopreserved testicular spermatozoa from the same nine patients and comparing all ICSI cycles performed; with fresh (25 cycles) and thawed (14 cycles) testicular spermatozoa, respectively: two-pronuclear fertilization, 47% versus 44%; embryo cleavage rates, 94% versus 89%; implantation rates, 9% versus 11%; and clinical PR, 26% versus 27%. The delivery or ongoing PR using fresh sperm was better (21% versus 9%), but the difference did not reach statistical significance. The cumulative clinical PRs and ongoing PRs per testicular sperm extraction procedure were 36% and 24%, respectively. CONCLUSION(S): Testicular sperm cryopreservation using a simple freezing protocol is promising in patients with nonobstructive azoospermia augmenting the overall success achieved after surgical sperm retrieval.

Testicular sperm retrieval by percutaneous fine needle sperm aspiration compared with testicular sperm extraction by open biopsy in men with non-obstructive azoospermia.

The efficiency of testicular sperm retrieval by testicular fine needle aspiration (TEFNA) was compared with open biopsy and testicular sperm extraction (TESE), in 37 rigorously selected patients with non-obstructive azoospermia. All patients underwent TEFNA and TESE consecutively. Thus, each patient served as his own control. The case was regarded as successful if at least one testicular spermatozoon was found allowing intracytoplasmic sperm injection (ICSI) of at least one oocyte. The mean age of the male patients was 32.7 years (range 24-47). Whereas by TEFNA spermatozoa enabling performance of ICSI were found in only four patients out of 37 (11%), open biopsy and TESE yielded spermatozoa in 16 cases (43%). The negative predictive value of high serum follicle stimulating hormone (FSH) concentrations (> or =10 IU/l) (predicting failure to find spermatozoa for ICSI) was low (38.4%). The positive predictive value (predicting the chance to find spermatozoa for ICSI) of normal-sized testicle was not different from that of small-sized (<15 ml) testicle (50%). Complications included one case of testicular bleeding following fine needle aspiration, treated locally, and two cases of extratunical haematomata following TESE requiring no intervention. In patients with non-obstructive azoospermia, TEFNA has a significantly lower yield compared to TESE. Performance of ICSI with testicular sperm in these cases resulted in satisfactory fertilization and high embryo transfer rates. The implantation and pregnancy rates per embryo transfer were 13 and 29% respectively. Neither serum FSH values nor testicular size were predictive of the chances to find spermatozoa for ICSI. Some complications may occur even following TEFNA.

Extended sperm preparation: an alternative to testicular sperm extraction in non-obstructive azoospermia.

Testicular sperm retrieval for the treatment of non-obstructive azoospermia requires the execution of an invasive procedure, with all its possible attending complications and subsequent long-term effects. This study suggests a new non-invasive approach for collection of spermatozoa in these patients: the extended sperm preparation (ESP). ESP consists of conducting a thorough microscopic search through many droplets of ejaculate sediment. ESP was performed for 49 patients; in 17 patients (35%), spermatozoa were found and subsequently used in intracytoplasmic sperm injection (ICSI). Of these preparations, five yielded fewer motile spermatozoa than the number of corresponding oocytes available, and in one patient only non-motile spermatozoa were recovered. The remaining 32 ESP-negative patients underwent testicular sperm extraction (TESE) from testicular biopsy. Spermatozoa were found in 16 of 32 biopsies (50%) and subsequently used in ICSI. Fertilization and cleavage rates were comparable in both ESP and TESE groups, yielding four clinical pregnancies in each group (27 and 29% respectively). Embryo morphology was defined as excellent in significantly more cases in the ESP group than the TESE group, and implantation rate appeared somewhat higher in the ESP group (16%) than the TESE group (13%). The ESP technique yields results similar to TESE, and can be applied in cases of non-obstructive azoospermia as a prerequisite modality enabling us to avoid testicular biopsy in 35% of cases.

Evaluation of chromatin condensation in human spermatozoa: a flow cytometric assay using acridine orange staining.

The quality of sperm chromatin is an important factor in fertilization and is especially critical where one spermatozoon is artificially selected for fertilizing an egg (as in intracytoplasmic sperm injection). In this study, flow cytometry after staining of human spermatozoa with Acridine Orange was used to study chromatin structure. A method is described for estimating the percentage of cells in a human sperm sample that have completed epididymal maturation in regard to chromatin condensation. Of the 121 samples of the semen that were examined, nine contained a higher percentage of hypocondensed spermatozoa and six samples contained elevated amounts of hypercondensed spermatozoa. In addition to aberrancies in chromatin condensation other defects showed up as satellite populations of spermatozoa with higher than normal ratios of red/green fluorescence after Acridine Orange staining. Such defects were found in 15 semen samples. The use of swim-up and Percoll gradient centrifugation methods was shown to improve the percentage of spermatozoa with normal chromatin structure in some samples with poor initial quality.