scholarly journals Complexin-I-deficient sperm are subfertile due to a defect in zona pellucida penetration

Reproduction ◽  
2008 ◽  
Vol 136 (3) ◽  
pp. 323-334 ◽  
Author(s):  
Longmei Zhao ◽  
Kerstin Reim ◽  
David J Miller
Keyword(s):  
Zona Pellucida ◽  
Semen Analysis ◽  
Fertilization Rate ◽  
Computer Assisted ◽  
Cell Fractionation ◽  
Synaptic Vesicle Exocytosis ◽  
Protein Receptor ◽  
Vesicle Exocytosis ◽  
Deficient Mice

Upon adhesion to the zona pellucida, sperm undergo regulated exocytosis of the acrosome. Although it is necessary for sperm to penetrate the zona pellucida and fertilize an egg, the acrosomal membrane fusion process is poorly understood. Complexins I and II are small, cytosolic proteins that bind to a complex of proteins termed the solubleN-ethylmaleimide-sensitive factor attachment protein receptor complex to regulate synaptic vesicle exocytosis. Complexin-II-deficient mice are fertile but the fertility of sperm from complexin-I-deficient male mice is unclear because the mice have ataxia and cannot mate. Here, we show that the genes encoding complexins I and II are expressed in primary spermatocytes and spermatids. Complexin proteins were found in/near the developing acrosome in spermatids and in or around the acrosome of mature sperm. Cell fractionation demonstrated that complexins I and II were predominantly found in the cytosolic fraction. Furthermore, sperm from complexin-I-deficient mice had normal morphology, number, and only small differences in motility, as assessed by computer-assisted semen analysis. Complexin-I-deficient sperm capacitated normally and bound to the zona pellucida. But when sperm from complexin-I-deficient mice were inseminated into females, a defect in fertility was observed, in concordance with previous data showing thatin vitrofertilization rate was also reduced. If the zona pellucida was removed prior toin vitrofertilization, fertility was normal, demonstrating that zona pellucida penetration was defective, a step requiring acrosomal exocytosis. Therefore, complexin-I-deficient sperm are subfertile due to faulty zona pellucida penetration.

163 HYPERACTIVATION OF STALLION SPERM IN FOLLICULAR FLUID FOR IN VITRO FERTILIZATION OF EQUINE OOCYTES

2012 ◽  
Vol 24 (1) ◽  
pp. 193 ◽  
Author(s):  
A. Lange-Consiglio ◽  
F. Cremonesi
Keyword(s):  
In Vitro Fertilization ◽  
Sperm Motility ◽  
Follicular Fluid ◽  
Zona Pellucida ◽  
Semen Analysis ◽  
Computer Assisted ◽  
Vitro Fertilization ◽  
Sperm Penetration ◽  
Low Incidence

In vitro fertilization has remained elusive in the horse, as evidenced by low sperm penetration rates when IVF has been attempted with in vivo- or in vitro-matured oocytes. It is likely that the low sperm penetration rates observed in IVF studies are due to the inability to appropriately capacitate or hyperactivate, or both, stallion sperm in the laboratory. The acquisition of hyperactivated sperm motility has been observed within the oviducts of mammals at the time of fertilization and is required for zona pellucida penetration in conjunction with the acrosome reaction (AR). Although the zona pellucida is considered the prime physiological inducer of AR, previous studies have shown a low incidence of AR in zona pellucida-bound stallion spermatozoa after 1 h of in vitro binding. This low incidence suggests that, besides the zona pellucida glycoproteins, another major factor might be responsible for AR. Protein-bound progesterone, present in equine follicular fluid (FF), has been demonstrated to induce AR in stallion spermatozoa. In this context, the aims of this study were (1) to hyperactivate stallion sperm in FF and (2) to verify whether this hyperactivation supports equine IVF. Pooled FF, aspirated from the preovulatory follicles of oestrous mares, was used and its progesterone concentration was determined by immune enzymatic assay. Spermatozoa from fertile stallions selected by a swim-up procedure were pre-incubated for 6 h in capacitating medium (modifed Whittens's medium (WM) supplemented with 25 mM NaHCO3 and 7 mg mL–1 of BSA) and then incubated for 6 h at 37°C in either FF or capacitating WM. Sperm motility was assayed by computer-assisted semen analysis, rates of AR were assessed by fluorescein isothiocyanate-PNA staining and rate of apoptosis was assessed by an annexin V test. For IVF, spermatozoa were incubated at 10 × 106 sperm mL–1 in capacitating WM for 6 h and then diluted to 1 × 106 sperm mL–1 in capacitating WM with or without 10% of FF. Five mature mare oocytes were transferred into droplets (100 μL) of the sperm suspensions covered with mineral oil and then incubated for 18 h at 38.5°C in 5% CO2 in humidified air. After that, oocytes were transferred to an embryo culture medium (DMEM/F-12) for an additional 3 days. Data were analysed by ANOVA. Treatment of sperm with FF resulted in a significant (P ≤ 0.05) decrease of 3 motility variables indicative of hyperactivation: straight line velocity, straightness and linearity. The highest rate of AR (29.44%) and a lower rate of apoptosis (16.93%) were obtained after 4 h of incubation in follicular fluid. By coupling capacitating conditions with the induction of hyperactivation using follicular fluid, we have obtained reproducible percentages of 8-cell-stage embryos (18.56%) in our IVF experiments. Conversely, sperm incubated in capacitating conditions but not treated with FF did not fertilize (0%). It is concluded that mare FF does not impair sperm viability, stimulates equine sperm hyperactivation in vitro, induces the AR and supports equine IVF.

An Insight into Novel Sperm Cell Proteins as Bio-markers for Male Infertility: A Review

2021 ◽  
Vol 21 ◽  
Author(s):  
Naina Kumar ◽  
Namit Kant Singh
Keyword(s):  
Male Infertility ◽  
Sperm Motility ◽  
Zona Pellucida ◽  
English Language ◽  
Semen Analysis ◽  
Meta Analysis ◽  
Sperm Dna Damage ◽  
Sperm Proteins

: Male infertility is rising now-a-days and accounts for major part of infertility cases worldwide. Novel tests are being developed for better detection and management of male infertility. Though there are many tests available for diagnosing male infertility like acrosome reaction rate, hemizona assay, in vivo or in vitro sperm penetration assay, sperm DNA damage tests, but semen analysis is most commonly used initial test for male infertility. It is usually associated with failure to detect cause in many cases, as seminal composition gets affected by a number of factors and can give false reports. Furthermore, it does not give any information about defects in capacitation, sperm Zona Pellucida interaction and sperm’s ability to fertilize oocytes. This results in failure of detection and delayed management of male infertility. Hence, the present review was conducted to identify various sperm proteins that play significant role in spermatogenesis, sperm motility, sperm-Zona Pellucida interaction and fertilization. These proteins can be used in future as markers of male infertility and will aid in better detection and management of male infertility. Methodology: Search for literature was made from 1970 to 2020 from various databases like PUBMED, SCOPUS, Google Scholar on sperm proteins and their role in male fertility using keywords: “sperm protein as bio-markers”, “novel sperm proteins as markers of infertility”, “Sperm proteins essential for capacitation, sperm motility and oocyte fertilization”. Inclusion criteria: All full-length research articles, systematic reviews, meta-analysis or abstracts on sperm proteins and male infertility published in English language in peer-reviewed journals were considered.

137 Assessment of fertilizing ability of Merino ram semen cold stored up to 48h by heterologous IVF of bovine oocytes

2019 ◽  
Vol 31 (1) ◽  
pp. 194 ◽  
Author(s):  
D. A. Galarza ◽  
M. Ladrón de Guevara ◽  
P. Beltrán-Breña ◽  
M. J. Sánchez-Calabuig ◽  
A. López-Sebastián ◽  
...  
Keyword(s):  
Semen Analysis ◽  
Skim Milk ◽  
Semen Parameters ◽  
Computer Assisted ◽  
Cleavage Rate ◽  
Straight Line ◽  
Fertilizing Ability ◽  
Pronuclear Formation ◽  
Bovine Oocytes

The use of cold-stored ram semen has been applied in sheep AI programs, because it preserves its fertilizing ability similar to fresh. Besides, the heterologous IVF has been successfully employed to assess semen fertilizing ability in several species. Hence, we aimed to evaluate the fertilizing ability of ram semen cold stored up to 48h at 5°C by assessing heterologous IVF using bovine oocytes. Fifteen pools of 3 normospermic Merino ram (2-7 years) ejaculates were collected using artificial vagina, diluted to 200×106 spermatozoa mL−1 with ultra-heat-treatment-based extender (skim milk-6% egg yolk) and cold stored up to 48h. In vitro matured zona-intact bovine oocytes were subjected to heterologous IVF using fresh semen (FS, n=707), semen cold stored to 24h (CS24, n=832) or semen cold stored to 48h (CS48, n=611). In parallel, homologous IVF (control, n=1356) and parthenogenesis (parth control non-fertilized oocytes, n=334) were performed. Ram non-selected and selected (BoviPure, Nidacon International, Mölndal, Sweden) semen parameters were evaluated by computer-assisted semen analysis. Sperm-oocyte interaction was assessed at 2.5h post-insemination (hpi) by evaluating the number of bound spermatozoa, whereas penetration and polyspermy were evaluated after 12 hpi. Presumptive zygotes were fixed and stained with Hoechst 33342 at 18, 20, 22, 24 and 26 hpi to assess pronuclear formation using phase contrast and confocal microscopy. Cleavage rate was evaluated in all groups at 48 hpi. Data obtained from 5 replicates were analysed using one-way ANOVA. Data was expressed as mean±standard error of the mean. In terms of sperm storage time, non-selected semen showed a significant decrease (P<0.05) for CS24 and CS48 compared with FS on progressive motility [SPM (%): 52.30±4.1 and 36.9±5.5v. 71.3±1.6] and straight-line velocity (mm s−1: 132.2±6.1 and 109.7±6.3v. 176.7±4.3), respectively. However, selected semen showed a decrease (P<0.05) only for CS48 when compared with CS24 or FS on SPM (35.6±3.9v. 56.1±6.91 and 59.3±2.6) and straight-line velocity (83.5±4.4v. 105.3±6.5 and 110±2.0), respectively. No differences were observed between heterologous IVF groups in all parameters evaluated. Homologous IVF showed a higher percentage of penetration only when compared with heterologous FS group (44.4±6.8v.12.5±4.5%; P<0.01). The polyspermy was higher in heterologous CS24 group when compared with homologous IVF (11.4±3.4v. 3.8±2.2; P<0.05). The homologous IVF group, as expected, showed the higher percentage of pronuclear formation at 18 hpi compared with heterologous IVF with FS (67.3±5.8v. 35.2±5.6%), CS24 (72.1±4.5 v. 37.2±5.7%) and CS48 (63.0±6.0 v. 27.0±5.6%), respectively (P<0.001). Likewise, cleavage rate was higher in homologous group compared with heterologous IVF and parthenogenetic groups for FS (78.3±2.6.8v. 46.3±3.2 and 7.0±2.3%), CS24 (78.4±2.6 v. 48.3±3.2 and 4.9±2.0%), and CS48 (78.4±3.3 v. 43.3±3.5 and 4.3±1.2%), respectively (P<0.001). In conclusion, Merino ram semen cold stored up to 48h maintains its fertilization ability to the same extend as fresh and can be used for sheep crossbreeding programs.

scholarly journals 12PREDICTION OF BULL FERTILITY BY COMPUTER ASSISTED SEMEN ANALYSIS

2004 ◽  
Vol 16 (2) ◽  
pp. 128 ◽  
Author(s):  
S. Cseh ◽  
T. Polichronopoulos ◽  
L. Solti
Keyword(s):  
Sperm Motility ◽  
Semen Analysis ◽  
Computer Assisted ◽  
Reproductive Process ◽  
Return Rates ◽  
Frozen Semen ◽  
Chi Square Analysis ◽  
Fertilizing Capability ◽  
Weighted Values

Sperm motility is clearly essential for fertilization both in vivo and in vitro. Motility is necessary for successful sperm transport, a step that is bypassed with in vitro fertilization. Recently, increasing attention has been paid to the objective evaluation and characterization of sperm motility more than simply determining the total proportion of motile spermatozoa. The purpose of computerassisted semen analysis (CASA) is to provide values for sperm concentration and sperm motility more rapidly and accurately than those obtained with traditional semen analyses methods. The objective of our experiment was to investigate the effect of specific aspects of sperm movement, such as the velocity of progression and the actual pattern of movement, to the fertilizing capability of sperm. Frozen semen samples of 10 HF breeding bulls were used in the study. For the motility analyses, Medealab CASA system (Medealab, Germany, Ver. 4.1) was used, and the velocity parameter of VCL (curvalinear velocity, μms−1), VSL (straight line velocity, μms−1), and VAP (average path velocity, μms−1) were evaluated and compared with the Day 30 and 75 non−return rates (NR30 and NR75). For every sample, a total of 10 fields were examined for 8s using a disposable 20 micron capillary chamber (CellVision, USA) giving a total of 1165 to 2831 cells evaluated. Chi square analysis, analyses of variance and linear correlation coefficient was applied to the statistical evaluation and comparison of the results. Data are based on weighted values. From the same batch of the analyzed frozen semen, a total of 8099 females were inseminated in more than 100 farms with a total of 6590 animals being positive for pregnancy at Day 30 and 4525 animals at Day 75. Within the bulls, differences were found in the values of NR30 and NR75 (P<0.05). Our data indicate very strong differences between the males’ NR30 and NR75 values (NR30: 65.6%±13.04 to 79.6%±11.17; P<0.001 and NR75: 37.8%±10.38 to 58.3%±15.53; P<0.001) reflecting the individual differences in the fertilizing capability of the males. All velocity parameters show very high correlation with strong significance both non−return rates but the best values belong to VAP (NR30 and NR75; P<0.02). Our data indicate that the bulls with lower VCL (25.51±33.04 to 79.54±58.03), VSL (11.35±19.45 to 36.36±35.71), and VAP (12.67±19.06 to 41.75±34.45) values showed lower fertilization rates both at NR30 and NR75. Computer and video technologies have advanced rapidly in recent years; thus the capability and accuracy of the latest versions of CASA systems are considerably better and they give more information about the different motion characteristics of spermatozoa. Because of the vital role of sperm motility in the reproductive process, such systems will enable us to move into a new era of diagnostic andrology and predict the fertilizing capability of semen. Supported by NKFP-Grants 4/040/2001 and 4/031/2001.

Effect of Panax notoginseng Extracts on Inferior Sperm Motility in vitro

1999 ◽  
Vol 27 (01) ◽  
pp. 123-128 ◽  
Author(s):  
Jung-Chou Chen ◽  
Ming-Xiong Xu ◽  
Leih-Der Chen ◽  
Yan-Nian Chen ◽  
Tsan Hung Chiu
Keyword(s):  
Sperm Motility ◽  
Sperm Count ◽  
Semen Analysis ◽  
Panax Notoginseng ◽  
World Health ◽  
Computer Assisted ◽  
Butanol Extract ◽  
The World ◽  
Health Organization

The purpose of this study was to investigate the effects of Panax notoginseng extracts on inferior sperm motility in vitro. Semen samples were collected from 23 patients with sperm motility between 20% and 40%. The sperm count was over 20 × 106/ml in accordance with the World Health Organization standard. 1.0 mg/ml and 2.0 mg/ml of Panax notoginseng extracts including aqueous extract, n-butanol extract, and polysaccharide fraction on sperm motility and progression were evaluated by computer assisted semen analysis. The results demonstrated that sperm motility as well as progression on inferior sperm motility were enhanced at 1 hour and 2 hours after incubation with all three types of extracts.

scholarly journals Effect of immunization of hamsters against recombinant P26h on fertility rates

Reproduction ◽  
2002 ◽  
pp. 307-313 ◽  
Author(s):  
C Gaudreault ◽  
L Montfort ◽  
R Sullivan
Keyword(s):  
Zona Pellucida ◽  
Binding Protein ◽  
Maltose Binding Protein ◽  
Fertilization Rate ◽  
Active Immunization ◽  
Antigenic Determinants ◽  
Binding Assays ◽  
Contraceptive Vaccine

Despite the various contraceptive methods available, an effective and inexpensive method remains to be established. Immunocontraception may help to achieve this goal. P26h has been proposed as a candidate for the development of a male contraceptive vaccine. P26h, a hamster sperm protein, interacts with the zona pellucida. Furthermore, in vivo fertilization can be blocked completely by active immunization of male hamsters against P26h. Maltose binding protein (MBP)-P26 shares antigenic determinants with the native P26h present on cauda epididymal spermatozoa. The aim of the present study was to reproduce the immunocontraceptive properties of native P26h by immunizing male hamsters against a recombinant P26h fused with a maltose binding protein (MBP). Active immunization of male hamsters with the MBP-P26h showed that specific anti-P26h circulating IgGs could be generated. Mating of immunized male hamsters with superovulated females resulted in a significant decrease, 20-25%, in the fertilization rate. This result is in agreement with results from in vitro sperm-zona pellucida binding assays. Indeed, the anti-recombinant P26h IgGs showed lower inhibitory properties when compared with anti-native P26h IgG. Despite the high anti-P26h IgG titres generated in hamsters, histological studies showed that active immunization has no pathological sequelae to the reproductive tissues. The potential of P26h as a candidate for a contraceptive vaccine is discussed.

scholarly journals Asthenozoospermia in Mice with Targeted Deletion of the Sperm Mitochondrion-Associated Cysteine-Rich Protein (Smcp) Gene

2002 ◽  
Vol 22 (9) ◽  
pp. 3046-3052 ◽  
Author(s):  
Karim Nayernia ◽  
Ibrahim M. Adham ◽  
Elke Burkhardt-Göttges ◽  
Jürgen Neesen ◽  
Mandy Rieche ◽  
...  
Keyword(s):  
Semen Analysis ◽  
Computer Assisted ◽  
Microscopical Examination ◽  
Structural Protein ◽  
Wild Type ◽  
Vitro Fertilization ◽  
In Vivo Experiments ◽  
Cysteine Rich Protein

ABSTRACT The sperm mitochondria-associated cysteine-rich protein (SMCP) is a cysteine- and proline-rich structural protein that is closely associated with the keratinous capsules of sperm mitochondria in the mitochondrial sheath surrounding the outer dense fibers and axoneme. To investigate the function of SMCP, we generated mice with a targeted disruption of the gene Smcp by homologous recombination. Homozygous mutant males on a mixed genetic background (C57BL/6J × 129/Sv) are fully fertile, while they are infertile on the 129/Sv background, although spermatogenesis and mating are normal. Homozygous Smcp−/− female mice are fertile on both genetic backgrounds. Electron microscopical examination demonstrated normal structures of sperm head, mitochondria, and tail. In vivo experiments with sperm of Smcp−/− 129/Sv mice revealed that the migration of spermatozoa from the uterus into the oviduct is reduced. This result is supported by the observation that sperm motility as determined by the computer-assisted semen analysis system (CASA) is significantly affected as compared to wild-type spermatozoa. In vitro fertilization assays showed that Smcp-deficient spermatozoa are able to bind to the oocyte but that the number of fertilized eggs is reduced by more than threefold relative to the wild-type control. However, removal of the zona pellucida resulted in an unaffected sperm-egg fusion which was monitored by the presence of pronuclei and generation of blastocyts. These results indicate that the infertility of the male Smcp−/− mice on the 129/Sv background is due to reduced motility of the spermatozoa and decreased capability of the spermatozoa to penetrate oocytes.

scholarly journals Inositol pyrophosphates inhibit synaptotagmin-dependent exocytosis

2016 ◽  
Vol 113 (29) ◽  
pp. 8314-8319 ◽  
Author(s):  
Tae-Sun Lee ◽  
Joo-Young Lee ◽  
Jae Won Kyung ◽  
Yoosoo Yang ◽  
Seung Ju Park ◽  
...  
Keyword(s):  
Synaptic Vesicle ◽  
Neurotransmitter Release ◽  
Hippocampal Neurons ◽  
Vesicle Fusion ◽  
Synaptic Membrane ◽  
Synaptic Vesicle Exocytosis ◽  
Dependent Inhibition ◽  
Vesicle Exocytosis ◽  
Inositol Pyrophosphates

Inositol pyrophosphates such as 5-diphosphoinositol pentakisphosphate (5-IP7) are highly energetic inositol metabolites containing phosphoanhydride bonds. Although inositol pyrophosphates are known to regulate various biological events, including growth, survival, and metabolism, the molecular sites of 5-IP7 action in vesicle trafficking have remained largely elusive. We report here that elevated 5-IP7 levels, caused by overexpression of inositol hexakisphosphate (IP6) kinase 1 (IP6K1), suppressed depolarization-induced neurotransmitter release from PC12 cells. Conversely, IP6K1 depletion decreased intracellular 5-IP7 concentrations, leading to increased neurotransmitter release. Consistently, knockdown of IP6K1 in cultured hippocampal neurons augmented action potential-driven synaptic vesicle exocytosis at synapses. Using a FRET-based in vitro vesicle fusion assay, we found that 5-IP7, but not 1-IP7, exhibited significantly higher inhibitory activity toward synaptic vesicle exocytosis than IP6. Synaptotagmin 1 (Syt1), a Ca2+ sensor essential for synaptic membrane fusion, was identified as a molecular target of 5-IP7. Notably, 5-IP7 showed a 45-fold higher binding affinity for Syt1 compared with IP6. In addition, 5-IP7–dependent inhibition of synaptic vesicle fusion was abolished by increasing Ca2+ levels. Thus, 5-IP7 appears to act through Syt1 binding to interfere with the fusogenic activity of Ca2+. These findings reveal a role of 5-IP7 as a potent inhibitor of Syt1 in controlling the synaptic exocytotic pathway and expand our understanding of the signaling mechanisms of inositol pyrophosphates.

scholarly journals A Review on Animals Semen Characteristics: Fertility, Reproduction and Development

2019 ◽  
pp. 1-9
Author(s):  
Farzad Moradpour
Keyword(s):  
Structural Integrity ◽  
Current Knowledge ◽  
Semen Analysis ◽  
Kinetic Property ◽  
Genetic Material ◽  
Computer Assisted ◽  
Semen Characteristics ◽  
The Individual

In this research, the goal of review was summarizing the current knowledge of the methods available to assess in vitro quality of frozen-thawed bovine spermatozoa also, a review on animal’s semen characteristics: fertility, reproduction and development after AI with that semen. Artificial insemination (AI) is the first generation reproductive biotechnology that has made a deep contribution to the genetics improvement in several animals. A fertile ejaculate must meet certain semen characteristics quality standards, such as: normal morphology, active energy metabolism, progressive motility, structural integrity and functionality of the membrane, penetration capacity and optimum transfer of genetic material. The percentage of total motile spermatozoa in normal canine ejaculates is between 70 to 90%. By the way, there are a lot of parameters that able to change on the composition and structure of various sperm plasma member domains, such as change temperature and sensitive to any theirs environments in vivo and vitro (tropical climates), season also nutrition. Computer-assisted semen analysis (CASA) is primarily used to obtain accurate and objective kinetic sperm measurements that gives extensive information about the kinetic property of the ejaculate based on measurements of the individual sperm cells.

scholarly journals Re-evaluation of the value of sperm morphology in classical in vitro fertilization in a Northeastern Chinese population

2019 ◽  
Vol 47 (9) ◽  
pp. 4134-4142 ◽  
Author(s):  
Dong-liang Zhu ◽  
Hong-guo Zhang ◽  
Rui-xue Wang ◽  
Yu-ting Jiang ◽  
Rui-zhi Liu
Keyword(s):  
In Vitro Fertilization ◽  
Chinese Population ◽  
Sperm Morphology ◽  
Semen Analysis ◽  
Fertilization Rate ◽  
Vitro Fertilization ◽  
Group 2 ◽  
Normal Fertilization ◽  
Group 1

Objective This study aimed to re-evaluate the clinical value of a 4% cut-off threshold of sperm morphology in in vitro fertilization (IVF) in a cohort of a Northeastern Chinese population. Methods A total of 375 IVF cycles that met strict inclusion criteria were included. These cycles were conducted with semen analysis and oocyte fertilization. A total of 188 embryo-transferred cycles proceeded. According to sperm morphology, 375 cycles were divided into group 1 (329 cycles, <4% normal sperm morphology rate [NSMR]) and group 2 (46 cycles, ≥4% NSMR), and 188 transferred cycles into group A (151 cycles, < 4% NSMR) and group B (37 cycles, ≥4% NSMR). Results The fertilization and normal fertilization rates were significantly lower in group 1 than in group 2. The normal fertilization rate was significantly correlated with an NSMR < 4% or ≥4%, but the fertilization rate was not significantly correlated with the NSMR. No significant differences were found in pregnancy outcomes between groups A and B. Conclusions This study suggests that infertile patients with an NSMR < 4% are more likely to have a poor normal fertilization status in IVF.

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