The latest WHO recommendations,2010 are based on semen parameters from approximately 2000 fertile men, from eight countries and three continents, whose partners achieved pregnancy within 12 months of unprotected sexual intercourse.
Pitfalls- huge shift in the lower reference values, one sided criteria.
Reference limits shouldn’t be over-interpreted
Interpret along with clinical history and physical examination.
Oration delivered by Dr Sujoy Dasgupta at Yuvacon, conference organized by the BOGS (Bengal Obstetric and Gynaecological Society) held on 22-23 April, 2023
The Need of LH in ART and Differences Between Sources of LH ActivitySandro Esteves
This document discusses the role of luteinizing hormone (LH) in reproductive cycles and ovarian stimulation. It summarizes that LH is essential for normal ovarian steroidogenesis and follicular development. Certain patient groups, such as older women, poor responders, and those with less sensitive ovaries may benefit from LH supplementation during controlled ovarian stimulation cycles to help maximize pregnancy rates. The document reviews several randomized controlled trials comparing recombinant follicle-stimulating hormone alone versus in combination with recombinant LH, finding improved outcomes with LH addition in some patient populations.
This document discusses reasons why IVF cycles may fail and provides guidance on learning from failed cycles. It defines recurrent IVF failure and recurrent implantation failure. Common causes of failure discussed include embryo quality, endometrial factors, and uterine issues like polyps or hydrosalpinx. Investigations like hysteroscopy and salpingectomy are recommended to address correctable causes. Other potential factors explored are endometrial thickness, scratching, and refractory endometrium. The goal is to identify avoidable causes and improve outcomes in subsequent cycles.
This document provides information on oocyte retrieval techniques and physics. It discusses the history of oocyte retrieval, moving from initial laparotomy techniques to modern laparoscopic and ultrasound-guided methods. It describes factors that affect oocyte collection such as pump vacuum, needle size and collection techniques. Potential complications of oocyte pickup including vaginal bleeding, intraperitoneal bleeding, and infection are also summarized. The document concludes by congratulating the authors on 35 years in the field of assisted reproductive technology.
This document provides information about semen analysis testing performed by the Department of Urology at Govt Royapettah Hospital and Kilpauk Medical College in Chennai, India. It lists the professors and assistant professors who moderate the tests and provides details about the purpose of semen analysis, the parameters evaluated, and the standard procedures for sample collection, analysis, and interpretation according to WHO guidelines. Semen analysis is a key test for evaluating male fertility and involves examining the sample macroscopically and microscopically to assess factors like volume, viscosity, pH, sperm concentration, motility, morphology, and vitality.
This document discusses the debate around whether embryo transfer should occur at the cleavage stage (day 3) or blastocyst stage (day 5-6).
It summarizes a study that found transferring embryos at the blastocyst stage resulted in a significantly higher ongoing pregnancy rate compared to cleavage stage in patients aged 35 or older, but no difference was seen in younger patients. The cumulative ongoing pregnancy rate was also higher but not significantly for blastocyst transfers.
The document also discusses some potential risks of blastocyst culture and transfer, including increased rates of preterm birth and large babies. Some studies found higher risks of congenital anomalies and epigenetic issues with blastocyst transfers as well.
Overall,
Ms. Doel Bose Pande discusses various semen preparation methods and principles. Common techniques include simple wash, swim-up, and density gradient centrifugation. The choice of method depends on factors like semen quality, intended use, and practical considerations. Density gradient is best for separating motile sperm from debris but is more time-consuming than direct swim-up. The goal is to recover high-quality sperm with minimal processing time and damage. Practical issues like sample volume and number of patients may also influence the choice of preparation technique.
This document describes the sperm chromatin dispersion (SCD) test process and interpretation. The SCD test involves adding semen samples to agarose on a microscope slide, staining it, and examining under a microscope. Sperm with intact DNA will show halos of dispersed chromatin around the core, while fragmented sperm will not. The document outlines the steps of heating and cooling the agarose, adding and incubating the semen sample, washing and staining the slide, then examining under a microscope. Sperm are scored and the percentage of sperm with fragmented DNA (SDF%) is calculated, with lower percentages indicating better sperm DNA integrity.
Oration delivered by Dr Sujoy Dasgupta at Yuvacon, conference organized by the BOGS (Bengal Obstetric and Gynaecological Society) held on 22-23 April, 2023
The Need of LH in ART and Differences Between Sources of LH ActivitySandro Esteves
This document discusses the role of luteinizing hormone (LH) in reproductive cycles and ovarian stimulation. It summarizes that LH is essential for normal ovarian steroidogenesis and follicular development. Certain patient groups, such as older women, poor responders, and those with less sensitive ovaries may benefit from LH supplementation during controlled ovarian stimulation cycles to help maximize pregnancy rates. The document reviews several randomized controlled trials comparing recombinant follicle-stimulating hormone alone versus in combination with recombinant LH, finding improved outcomes with LH addition in some patient populations.
This document discusses reasons why IVF cycles may fail and provides guidance on learning from failed cycles. It defines recurrent IVF failure and recurrent implantation failure. Common causes of failure discussed include embryo quality, endometrial factors, and uterine issues like polyps or hydrosalpinx. Investigations like hysteroscopy and salpingectomy are recommended to address correctable causes. Other potential factors explored are endometrial thickness, scratching, and refractory endometrium. The goal is to identify avoidable causes and improve outcomes in subsequent cycles.
This document provides information on oocyte retrieval techniques and physics. It discusses the history of oocyte retrieval, moving from initial laparotomy techniques to modern laparoscopic and ultrasound-guided methods. It describes factors that affect oocyte collection such as pump vacuum, needle size and collection techniques. Potential complications of oocyte pickup including vaginal bleeding, intraperitoneal bleeding, and infection are also summarized. The document concludes by congratulating the authors on 35 years in the field of assisted reproductive technology.
This document provides information about semen analysis testing performed by the Department of Urology at Govt Royapettah Hospital and Kilpauk Medical College in Chennai, India. It lists the professors and assistant professors who moderate the tests and provides details about the purpose of semen analysis, the parameters evaluated, and the standard procedures for sample collection, analysis, and interpretation according to WHO guidelines. Semen analysis is a key test for evaluating male fertility and involves examining the sample macroscopically and microscopically to assess factors like volume, viscosity, pH, sperm concentration, motility, morphology, and vitality.
This document discusses the debate around whether embryo transfer should occur at the cleavage stage (day 3) or blastocyst stage (day 5-6).
It summarizes a study that found transferring embryos at the blastocyst stage resulted in a significantly higher ongoing pregnancy rate compared to cleavage stage in patients aged 35 or older, but no difference was seen in younger patients. The cumulative ongoing pregnancy rate was also higher but not significantly for blastocyst transfers.
The document also discusses some potential risks of blastocyst culture and transfer, including increased rates of preterm birth and large babies. Some studies found higher risks of congenital anomalies and epigenetic issues with blastocyst transfers as well.
Overall,
Ms. Doel Bose Pande discusses various semen preparation methods and principles. Common techniques include simple wash, swim-up, and density gradient centrifugation. The choice of method depends on factors like semen quality, intended use, and practical considerations. Density gradient is best for separating motile sperm from debris but is more time-consuming than direct swim-up. The goal is to recover high-quality sperm with minimal processing time and damage. Practical issues like sample volume and number of patients may also influence the choice of preparation technique.
This document describes the sperm chromatin dispersion (SCD) test process and interpretation. The SCD test involves adding semen samples to agarose on a microscope slide, staining it, and examining under a microscope. Sperm with intact DNA will show halos of dispersed chromatin around the core, while fragmented sperm will not. The document outlines the steps of heating and cooling the agarose, adding and incubating the semen sample, washing and staining the slide, then examining under a microscope. Sperm are scored and the percentage of sperm with fragmented DNA (SDF%) is calculated, with lower percentages indicating better sperm DNA integrity.
Sperm Function Tests are the keystones of evaluating functional condition of sperms. The fertility potential of a sperm will be decided not only with the number & motility but with the functional competence which is of utmost importance.
ICSI as it is presently performed is far from an ideal solution because the selection of sperm is based on the judgement of an embryologist, who is looking for the most normal appearing sperm available.
Anti Mullerian hormone -(AMH) -All Facts You Should Know | A Crucial Hormone ...martinshaji
AMH, or anti-Müllerian hormone, is a substance produced by granulosa cells in ovarian follicles. It is used as a biomarker for fertility potential. AMH levels can provide information about the number of follicles in a woman's ovaries and thus her egg reserve. An AMH test involves a blood draw to measure AMH levels. Higher AMH levels generally indicate better fertility and egg quality, while lower levels suggest reduced fertility and a smaller egg reserve. AMH testing can help assess fertility, the need for fertility treatments, and menopausal transition.
This document describes various ovarian stimulation protocols for infertility treatment, including oral medications, injectable medications, and monitoring techniques. It summarizes protocols for natural cycles, mild stimulation, conventional stimulation, antagonist protocols, and protocols for poor responders. Key points include the use of clomiphene citrate, gonadotropins like hMG and rFSH, protocols with and without downregulation, monitoring with ultrasound and hormones, and tailoring the protocol based on ovarian reserve and previous response. The goal is to recruit multiple follicles for retrieval while avoiding overstimulation and maintaining endometrial receptivity.
MANAGEMENT OF POOR RESPONDERS IN IVF BY DR SHASHWAT JANIDR SHASHWAT JANI
This document discusses the management of poor responders to ovarian stimulation. It defines poor responders according to the Bologna criteria as having two of the following: advanced age, a previous poor response, or abnormal biomarkers of ovarian reserve. It identifies various risk factors for poor response and stresses the importance of predicting response before treatment. It then discusses individualized controlled ovarian stimulation, including increasing gonadotropin doses, modifying GnRH analog protocols, using GnRH antagonists, and supplementing with growth hormone, estradiol, recombinant LH, and androgens to potentially improve outcomes for poor responders.
This document discusses the pros and cons of transferring embryos on day 5 (blastocyst stage) versus day 3. It raises questions about whether day 5 transfer should be routine practice and whether there are any adverse effects. Specifically, it notes that day 5 transfer is not suitable for all women, especially those with a limited number of embryos, and that an increased incidence of autism has been reported. It also discusses whether day 5 transfer is practical given the infrastructure needed, and whether it is really of any value if more than one embryo is being transferred. The conclusion is that day 5 transfer should only be offered for highly selected cases.
The document discusses the Sperm Chromatin Dispersion Test, which is used to assess DNA fragmentation in sperm. It works by using an acid treatment to denature fragmented sperm DNA, then removing nuclear proteins to allow dispersed DNA loops to form halos around nucleoids. For sperm with fragmented DNA, little to no halo is observed. Treatments discussed to address DNA fragmentation include antibiotics, antioxidants, lifestyle changes like quitting smoking, and medical procedures. The test is shown to be useful for diagnosing infertility issues and evaluating treatment effectiveness.
Novel treatments to trigger final follicular maturation and luteal phase supportSandro Esteves
This document summarizes novel strategies for triggering final follicular maturation and supporting the luteal phase in fertility treatments. It discusses evaluating the quality of trigger and luteal phase support methods based on indicators of safety, effectiveness, and patient-centeredness. Specific strategies used at Androfert clinic are presented, including individualizing triggers and support according to patient risk factors. Recombinant hCG is shown to have advantages over urinary hCG in terms of effectiveness, safety, and patient preferences. GnRH agonist triggering avoids risk of ovarian hyperstimulation syndrome but needs additional luteal phase support.
This document discusses various sperm preparation methods used for intrauterine insemination (IUI). It describes the simple wash method, swim-up method, and density gradient centrifugation method. For each method, it provides the steps, including centrifugation speeds and times. It also covers preparing samples for retrograde ejaculation and samples from HIV-infected patients. The goal of sperm preparation is to select motile sperm and remove other seminal constituents like debris to maximize the chances of fertilization during IUI.
Semen analysis as per WHO and clinical implicationsSandro Esteves
This document discusses semen analysis, new WHO reference values from 2010, and implications for clinical management. Key points include:
- Semen analysis provides biomarkers to predict fertility potential but has limitations on its own.
- The 2010 WHO reference values were derived from a meta-analysis of recent father studies and established new lower cut-off limits.
- The new WHO values resulted in many patients being reclassified from abnormal to normal ranges, especially for morphology, and impacted referral patterns, treatment recommendations, and access to ART.
- A comprehensive male fertility evaluation requires more than just a semen analysis and should include history, physical exam, hormones, and advanced sperm testing to properly diagnose and manage cases.
This document provides an overview of IVF and ICSI procedures. It discusses that IVF involves fertilizing eggs with sperm in a lab dish, then transferring embryos into the uterus. ICSI is used for severe male factor infertility and involves injecting a single sperm into each egg. Both aim to increase the chances of fertilization. The document outlines the steps of ovarian stimulation, egg retrieval, sperm preparation, fertilization, embryo culture, and embryo transfer.
Ovarian reserve refers to the reproductive potential left within a woman's two ovaries based on number and quality of eggs. Diminished ovarian reserve is the loss of normal reproductive potential in the ovaries due to a lower count or quality of the remaining eggs
This document discusses various sperm preparation techniques used prior to assisted reproductive technologies. It begins by explaining the reasons for processing sperm samples, such as removing components that could prevent pregnancy and selecting morphologically normal, motile sperm free of debris. Several migration-based techniques are described, including swim-up, density gradient centrifugation, and magnetic activated cell sorting. Glass wool filtration and zeta potential techniques are also covered. The document discusses preparation of epididymal and testicular sperm samples. It concludes by outlining methods for preparing sperm from retrograde ejaculation samples.
MACS is a sperm separation technique used in A.R.T. It allows separation of good quality sperm from apoptotic sperm (Sperm programmed to die) which may be one of the causes for failure of IVF
https://www.novaivifertility.com/primary-offering/andrology
Oocyte retrieval involves three key steps:
1) Anesthetizing the patient using either conscious sedation with a paracervical block or general anesthesia. 2) Guiding an ultrasound-monitored needle into each follicle to aspirate the follicular fluid and oocyte. 3) Using a suction pump set to 90-120 mmHg for mature follicles and 40-60 mmHg for immature follicles to aspirate the fluid and oocyte without damaging the cumulus-oocyte complex. Precautions like antibiotics and monitoring for bleeding are important to minimize risks of complications.
It is very important to refer proper patient at proper time for infertility treatment. This presentation explores briefly the different criteria to refer the patient and the follow-up after.
This document provides an overview of intrauterine insemination (IUI). Some key points include:
IUI is a first-line, non-invasive fertility treatment that involves placing processed sperm directly into the uterus. Success rates range from 6-20% depending on the stimulation protocol used. Factors like age, infertility duration and etiology, and semen quality impact success rates. Strict monitoring is important to minimize risks of ovarian hyperstimulation syndrome while maximizing pregnancy chances. Proper sperm processing techniques and timing of insemination relative to ovulation are also important considerations for IUI.
This document discusses poor ovarian responders in assisted reproductive technology treatment. It defines poor ovarian response based on the Bologna criteria of fewer than 3 oocytes retrieved with conventional stimulation. Poor responders make up 9-24% of IVF patients. The document outlines limitations of the Bologna criteria and introduces the POSEIDON classification system, which categorizes patients based on age, ovarian reserve markers, and ovarian response. It discusses markers for predicting poor response and reviews strategies for managing poor responders, including use of recombinant FSH, increasing FSH dosage, adding recombinant LH, and DHEA supplementation.
Empty follicle syndrome is a complication where mature follicles fail to yield oocytes during egg retrieval despite optimal stimulation and triggering. It can be genuine, with no clear cause, or false due to issues with the hCG trigger shot. Risk factors include advanced age, infertility duration, and low ovarian reserve. Treatment of false EFS involves re-administering hCG and repeating retrieval after 36 hours. Prognosis is generally good except for recurrent cases. Ongoing research aims to better understand the mechanisms and identify prevention strategies.
This document provides information about performing a semen analysis, including definitions of key terms, procedures, and normal ranges. It discusses:
1. The components and normal appearance of semen after liquefaction.
2. Procedures for assessing semen volume, viscosity, pH, sperm motility, morphology, concentration, and vitality under a microscope.
3. Factors that can affect the results, such as abstinence time, sample handling, and medical conditions.
The document is intended as a reference for medical professionals performing semen analyses to evaluate male fertility and identify potential issues.
The document provides information about andrology laboratory services for male infertility evaluation and treatment. It discusses:
- Tests offered including semen analysis, specialized tests of sperm function and morphology, sperm processing for infertility treatments, and cryopreservation.
- Procedures for semen sample collection, transport, and analysis following WHO standards, including macroscopic examination of volume, pH, and microscopic examination of motility, concentration, vitality, and morphology.
- Uses of semen analysis to diagnose infertility issues, identify treatment options, and assess effectiveness of treatments like vasectomy reversal. Computer-assisted semen analysis is also discussed.
Sperm Function Tests are the keystones of evaluating functional condition of sperms. The fertility potential of a sperm will be decided not only with the number & motility but with the functional competence which is of utmost importance.
ICSI as it is presently performed is far from an ideal solution because the selection of sperm is based on the judgement of an embryologist, who is looking for the most normal appearing sperm available.
Anti Mullerian hormone -(AMH) -All Facts You Should Know | A Crucial Hormone ...martinshaji
AMH, or anti-Müllerian hormone, is a substance produced by granulosa cells in ovarian follicles. It is used as a biomarker for fertility potential. AMH levels can provide information about the number of follicles in a woman's ovaries and thus her egg reserve. An AMH test involves a blood draw to measure AMH levels. Higher AMH levels generally indicate better fertility and egg quality, while lower levels suggest reduced fertility and a smaller egg reserve. AMH testing can help assess fertility, the need for fertility treatments, and menopausal transition.
This document describes various ovarian stimulation protocols for infertility treatment, including oral medications, injectable medications, and monitoring techniques. It summarizes protocols for natural cycles, mild stimulation, conventional stimulation, antagonist protocols, and protocols for poor responders. Key points include the use of clomiphene citrate, gonadotropins like hMG and rFSH, protocols with and without downregulation, monitoring with ultrasound and hormones, and tailoring the protocol based on ovarian reserve and previous response. The goal is to recruit multiple follicles for retrieval while avoiding overstimulation and maintaining endometrial receptivity.
MANAGEMENT OF POOR RESPONDERS IN IVF BY DR SHASHWAT JANIDR SHASHWAT JANI
This document discusses the management of poor responders to ovarian stimulation. It defines poor responders according to the Bologna criteria as having two of the following: advanced age, a previous poor response, or abnormal biomarkers of ovarian reserve. It identifies various risk factors for poor response and stresses the importance of predicting response before treatment. It then discusses individualized controlled ovarian stimulation, including increasing gonadotropin doses, modifying GnRH analog protocols, using GnRH antagonists, and supplementing with growth hormone, estradiol, recombinant LH, and androgens to potentially improve outcomes for poor responders.
This document discusses the pros and cons of transferring embryos on day 5 (blastocyst stage) versus day 3. It raises questions about whether day 5 transfer should be routine practice and whether there are any adverse effects. Specifically, it notes that day 5 transfer is not suitable for all women, especially those with a limited number of embryos, and that an increased incidence of autism has been reported. It also discusses whether day 5 transfer is practical given the infrastructure needed, and whether it is really of any value if more than one embryo is being transferred. The conclusion is that day 5 transfer should only be offered for highly selected cases.
The document discusses the Sperm Chromatin Dispersion Test, which is used to assess DNA fragmentation in sperm. It works by using an acid treatment to denature fragmented sperm DNA, then removing nuclear proteins to allow dispersed DNA loops to form halos around nucleoids. For sperm with fragmented DNA, little to no halo is observed. Treatments discussed to address DNA fragmentation include antibiotics, antioxidants, lifestyle changes like quitting smoking, and medical procedures. The test is shown to be useful for diagnosing infertility issues and evaluating treatment effectiveness.
Novel treatments to trigger final follicular maturation and luteal phase supportSandro Esteves
This document summarizes novel strategies for triggering final follicular maturation and supporting the luteal phase in fertility treatments. It discusses evaluating the quality of trigger and luteal phase support methods based on indicators of safety, effectiveness, and patient-centeredness. Specific strategies used at Androfert clinic are presented, including individualizing triggers and support according to patient risk factors. Recombinant hCG is shown to have advantages over urinary hCG in terms of effectiveness, safety, and patient preferences. GnRH agonist triggering avoids risk of ovarian hyperstimulation syndrome but needs additional luteal phase support.
This document discusses various sperm preparation methods used for intrauterine insemination (IUI). It describes the simple wash method, swim-up method, and density gradient centrifugation method. For each method, it provides the steps, including centrifugation speeds and times. It also covers preparing samples for retrograde ejaculation and samples from HIV-infected patients. The goal of sperm preparation is to select motile sperm and remove other seminal constituents like debris to maximize the chances of fertilization during IUI.
Semen analysis as per WHO and clinical implicationsSandro Esteves
This document discusses semen analysis, new WHO reference values from 2010, and implications for clinical management. Key points include:
- Semen analysis provides biomarkers to predict fertility potential but has limitations on its own.
- The 2010 WHO reference values were derived from a meta-analysis of recent father studies and established new lower cut-off limits.
- The new WHO values resulted in many patients being reclassified from abnormal to normal ranges, especially for morphology, and impacted referral patterns, treatment recommendations, and access to ART.
- A comprehensive male fertility evaluation requires more than just a semen analysis and should include history, physical exam, hormones, and advanced sperm testing to properly diagnose and manage cases.
This document provides an overview of IVF and ICSI procedures. It discusses that IVF involves fertilizing eggs with sperm in a lab dish, then transferring embryos into the uterus. ICSI is used for severe male factor infertility and involves injecting a single sperm into each egg. Both aim to increase the chances of fertilization. The document outlines the steps of ovarian stimulation, egg retrieval, sperm preparation, fertilization, embryo culture, and embryo transfer.
Ovarian reserve refers to the reproductive potential left within a woman's two ovaries based on number and quality of eggs. Diminished ovarian reserve is the loss of normal reproductive potential in the ovaries due to a lower count or quality of the remaining eggs
This document discusses various sperm preparation techniques used prior to assisted reproductive technologies. It begins by explaining the reasons for processing sperm samples, such as removing components that could prevent pregnancy and selecting morphologically normal, motile sperm free of debris. Several migration-based techniques are described, including swim-up, density gradient centrifugation, and magnetic activated cell sorting. Glass wool filtration and zeta potential techniques are also covered. The document discusses preparation of epididymal and testicular sperm samples. It concludes by outlining methods for preparing sperm from retrograde ejaculation samples.
MACS is a sperm separation technique used in A.R.T. It allows separation of good quality sperm from apoptotic sperm (Sperm programmed to die) which may be one of the causes for failure of IVF
https://www.novaivifertility.com/primary-offering/andrology
Oocyte retrieval involves three key steps:
1) Anesthetizing the patient using either conscious sedation with a paracervical block or general anesthesia. 2) Guiding an ultrasound-monitored needle into each follicle to aspirate the follicular fluid and oocyte. 3) Using a suction pump set to 90-120 mmHg for mature follicles and 40-60 mmHg for immature follicles to aspirate the fluid and oocyte without damaging the cumulus-oocyte complex. Precautions like antibiotics and monitoring for bleeding are important to minimize risks of complications.
It is very important to refer proper patient at proper time for infertility treatment. This presentation explores briefly the different criteria to refer the patient and the follow-up after.
This document provides an overview of intrauterine insemination (IUI). Some key points include:
IUI is a first-line, non-invasive fertility treatment that involves placing processed sperm directly into the uterus. Success rates range from 6-20% depending on the stimulation protocol used. Factors like age, infertility duration and etiology, and semen quality impact success rates. Strict monitoring is important to minimize risks of ovarian hyperstimulation syndrome while maximizing pregnancy chances. Proper sperm processing techniques and timing of insemination relative to ovulation are also important considerations for IUI.
This document discusses poor ovarian responders in assisted reproductive technology treatment. It defines poor ovarian response based on the Bologna criteria of fewer than 3 oocytes retrieved with conventional stimulation. Poor responders make up 9-24% of IVF patients. The document outlines limitations of the Bologna criteria and introduces the POSEIDON classification system, which categorizes patients based on age, ovarian reserve markers, and ovarian response. It discusses markers for predicting poor response and reviews strategies for managing poor responders, including use of recombinant FSH, increasing FSH dosage, adding recombinant LH, and DHEA supplementation.
Empty follicle syndrome is a complication where mature follicles fail to yield oocytes during egg retrieval despite optimal stimulation and triggering. It can be genuine, with no clear cause, or false due to issues with the hCG trigger shot. Risk factors include advanced age, infertility duration, and low ovarian reserve. Treatment of false EFS involves re-administering hCG and repeating retrieval after 36 hours. Prognosis is generally good except for recurrent cases. Ongoing research aims to better understand the mechanisms and identify prevention strategies.
This document provides information about performing a semen analysis, including definitions of key terms, procedures, and normal ranges. It discusses:
1. The components and normal appearance of semen after liquefaction.
2. Procedures for assessing semen volume, viscosity, pH, sperm motility, morphology, concentration, and vitality under a microscope.
3. Factors that can affect the results, such as abstinence time, sample handling, and medical conditions.
The document is intended as a reference for medical professionals performing semen analyses to evaluate male fertility and identify potential issues.
The document provides information about andrology laboratory services for male infertility evaluation and treatment. It discusses:
- Tests offered including semen analysis, specialized tests of sperm function and morphology, sperm processing for infertility treatments, and cryopreservation.
- Procedures for semen sample collection, transport, and analysis following WHO standards, including macroscopic examination of volume, pH, and microscopic examination of motility, concentration, vitality, and morphology.
- Uses of semen analysis to diagnose infertility issues, identify treatment options, and assess effectiveness of treatments like vasectomy reversal. Computer-assisted semen analysis is also discussed.
Semen analysis provides important information about male fertility. It involves examining semen volume, color, pH, sperm count, motility, morphology, and the presence of any abnormalities. The analysis evaluates the main components of semen from the testes, prostate, and other glands. It is used to assess infertility, check for issues after vasectomy, and for sperm donations. Proper sample collection and handling is important for accuracy of the results.
The document provides information about semen analysis, including the typical components and fractions of semen, the structures and sizes of sperm cells, indications for semen analysis, the process of sample collection and handling, and the steps involved in an initial semen analysis. Key points include that semen is made up of contributions from various glands, liquefies within 15-30 minutes, and an analysis involves examining the sample under a microscope to assess characteristics like motility, viscosity, and presence of other cells.
Semen analysis WHO 2010 BY DR ANAMIKA DEVAnamika Dev
The document provides information about semen analysis, including the typical components and fractions of semen, the structures and sizes of sperm cells, indications for semen analysis, the process of sample collection and handling, and the steps involved in an initial semen analysis. Key points include that semen is made up of contributions from various glands, liquefies within 15-30 minutes, and an analysis involves examining the sample under a microscope to assess characteristics like motility, viscosity, and presence of other cells.
Tolerance to tissue and cell antigens can be
induced by injection of hemopoietic (stem)
cells in neonatal or severely
immunocompromised (by lethal irradiation
or drug treatment) animals.
Also, grafting of allogeneic bone marrow or
thymus in early life results in tolerance to
the donor type cells and tissues. Such
animals are known as chimeras. These
findings are of significant practical
application in bone marrow grafting
This document summarizes histological and cytological specimens. It describes tissue samples obtained from biopsies or autopsies that can be either histological (muscular) or cytological (liquid). Cytological specimens include bodily fluids like urine, cerebrospinal fluid, joint fluid, endoscopy samples, sputum, and other fluids. The procedures for processing and staining samples of each fluid type are provided. Semen is also discussed as a cytological specimen, including normal parameters for analysis and staining techniques.
This is an important topic of mammalian (Male) reproductive toxicology.By doing this test sperm abnormalities should be cured. This topic is available in net but not like, what a master student try to find out.If there is anything wrong then correct me please.
In this ppt i have included methods of semen analysis and the importance and some agents which create semen abnormalities.
This document provides information about performing a semen analysis. It discusses the structures involved in semen production, contributions to semen volume, indications for a semen analysis, how to collect a semen sample, and the various tests that can be done on a semen sample including physical examination, microscopic examination of sperm count, motility, viability and morphology, immunologic analysis for antisperm antibodies, biochemical analysis of fructose, and sperm function tests. A normal semen analysis provides important information for investigating male factor infertility while abnormal results can help identify potential causes of reduced fertility.
Intrauterine insemination (IUI) is a fertility treatment that involves placing sperm inside a woman’s uterus to facilitate fertilisation. The goal of IUI is to increase the number of sperm that reach the fallopian tubes and subsequently increase the chance of fertilisation.
This document discusses semen analysis, which plays a key role in evaluating male infertility. It describes the normal parameters for semen volume, pH, liquefaction time, viscosity, sperm concentration, motility, morphology, and viability. The roles of the prostate, seminal vesicles, and other glands in contributing to the semen are outlined. Standard procedures for semen collection and analysis are provided, along with causes of abnormal results and limitations of semen analysis. Advanced computer-assisted techniques for further evaluating sperm motility and function are also mentioned.
A semen analysis measures the amount and quality of semen and sperm to determine fertility. It analyzes semen volume, pH, sperm count, motility, morphology, and vitality. Sperm are produced in the testes and transported through the seminal vesicles and prostate to provide nutrients and protection. A semen analysis is an important first test for infertility, identifying potential issues affecting over 1/3 of couples unable to conceive.
Semen contains spermatozoa and seminal plasma secreted from glands. Semen stains may be examined in criminal cases like rape. Samples are collected without heat and packaged carefully. Screening tests examine color, fluorescence under UV light, and chemical tests for choline or spermine. Confirmatory tests identify intact spermatozoa under microscopy or seminal markers like acid phosphatase. Motility can determine timing of deposition. Species origin is confirmed by precipitin tests, LDH patterns, or presence of human-specific Y bodies in sperm.
1. A semen analysis measures the amount and quality of semen and sperm to determine fertility. Problems with semen or sperm affect over 1/3 of infertile couples.
2. Semen is composed of sperm (2-5%), seminal fluid (60-75%), prostate fluid (25-30%), and bulbourethral gland fluid (1-5%). The sperm must be able to move and function within the fluid.
3. A semen analysis evaluates semen volume, viscosity, pH, sperm count, motility, and morphology. Normal ranges are provided for each parameter. Problems with any parameter can indicate infertility.
A semen analysis measures the amount and quality of semen and sperm. It is one of the first tests done to evaluate male fertility issues. It determines semen volume, sperm concentration, motility, morphology, pH, and presence of white blood cells or other abnormalities. The sample is collected through masturbation and analyzed under a microscope to measure these parameters. Normal ranges are defined by the WHO for factors like sperm concentration, motility, and morphology. Abnormal results can indicate issues that reduce fertility. The analysis provides important information about male reproductive health and ability to conceive.
A semen analysis measures the amount and quality of semen and sperm. It is one of the first tests done to evaluate male fertility issues. It determines semen volume, sperm concentration, motility, morphology, pH and presence of white blood cells or other abnormalities. The sample is collected through masturbation and analyzed under a microscope to measure these parameters. Normal ranges have been established by the WHO. Abnormal results could indicate issues that reduce fertility. The analysis provides important information about male reproductive health and ability to conceive.
Semen examination provides important evidence in sexual assault cases. Semen is composed of sperm cells suspended in seminal fluid. Initial presumptive tests for semen examine for the presence of enzymes like acid phosphatase. Confirmatory testing requires microscopic examination of the stain to identify sperm cells, as their presence confirms the source was human semen. Proper collection and handling of evidence from victims and suspects is crucial, as it allows laboratory examination to detect potential DNA transfers that can identify assailants. Semen evidence plays a key role in medico-legal cases involving rape, sodomy, incest, and disputed paternity.
A semen analysis measures the volume, pH, sperm count, motility, morphology, and other parameters of a semen sample. It is performed to evaluate male fertility and determine if issues such as infertility, a failed vasectomy, or failed vasectomy reversal are present. Proper collection involves abstaining from ejaculation for 2-5 days prior and collecting the sample by masturbation within 1 hour of the analysis. The semen is then examined under a microscope to evaluate the various sperm characteristics and identify any abnormalities.
The document provides guidelines for analyzing stool samples in a laboratory setting. Key steps include collecting the sample in a clean cup, examining a thin smear of the stool under the microscope using 10x and 40x lenses, and noting any observations of mucus, pus, blood cells, parasites, or other abnormal findings. The results are interpreted to help diagnose common gastrointestinal issues or parasites present in the stool. Multiple samples over several days may be needed to rule out the presence of parasites if not seen on initial examination.
Similar to sperm assessment- traditional and novel approaches.pptx (20)
Abortion Including Recurrent Abortion And Septic Abortion.pptxDeepekaTS
Abortion is defined as the spontaneous or induced termination of pregnancy
before fetal viability. Many prefer miscarriage for spontaneous loss.
abortion as
loss or termination of a pregnancy with a fetus aged younger than 20 weeks’
gestation or weighing <500 g.
Of all miscarriages, approximately half are euploid abortions, that is, carrying a normal chromosomal complement.
Most common abnormalities are
trisomy, found in 50 to 60 percent;
monosomy X, in 9 to 13 percent; and
triploidy, in 11 to 12 percent
A prominent miscarriage risk is associated with poorly
controlled diabetes mellitus, obesity, thyroid disease, and systemic lupus
erythematosus. In these, inflammatory mediators may be an underlying theme
to pregnancy loss.
For women undergoing cancer treatment, direct therapeutic radiation can
cause miscarriage.
Intraabdominal abcess- types and management .pptxDeepekaTS
This document discusses different types of intra-abdominal abscesses including their locations, symptoms, investigations, and management. It describes subphrenic, pelvic, and perinephric abscesses in more detail. Symptoms of intra-abdominal abscesses are generally vague but may include low grade fever, tachycardia, and localized tenderness. Investigations include blood tests showing leukocytosis as well as imaging like ultrasound, CT scan, and radiolabeled white blood cell scans. Management involves CT-guided or surgical drainage depending on the location and type of abscess. The document also discusses ascites including causes, clinical features, evaluation, treatment, and complications related to ascites.
Blood loss of >/ 500 ml within 24 hours of vaginal birth or 1000 ml after caesarean section or any blood loss sufficient to compromise haemodynamic instability
MINOR PPH- 500- 1000ml blood loss
MAJOR PPH- > 1000ml Blood loss
MASSIVE PPH- >2000ml Blood loss
Dystocia literally means difficult labor and is characterized by abnormally
slow labor progress.
DYSTOCIA
ABNORMALITIES OF THE EXPULSIVE FORCES
PREMATURELY RUPTURED MEMBRANES AT TERM
PRECIPITOUS LABOR AND DELIVERY
FETOPELVIC DISPROPORTION
FACE PRESENTATION
BROW PRESENTATION
TRANSVERSE LIE
COMPOUND PRESENTATION
COMPLICATIONS WITH DYSTOCIA
Chronic pelvic pain can be defined as intermittent or constant pain in the lower abdomen or pelvis of a woman of at least 6 months in duration, not occurring exclusively with menstruation or intercourse and not associated with pregnancy.
This document provides information on pelvic organ prolapse. It defines prolapse as the descent of genital organs through the pelvic floor. It describes the three levels of pelvic support and the types of prolapse that can occur at each level. Symptoms, signs, grading systems, etiology related to childbirth, and risk factors are outlined. Both conservative treatments like pelvic floor exercises and pessaries as well as various surgical repair options to correct prolapse in the anterior, posterior, and apical compartments are summarized.
This document provides guidelines for interpreting CTG traces during labor and delivery. It outlines appropriate monitoring frequencies and parameters to document. It also describes conservative measures that can be taken if traces show some abnormalities, as well as guidelines for fetal blood sampling. Cutoffs for normal, borderline, and abnormal pH and lactate levels are provided. Stages of labor are defined and appropriate monitoring and actions are outlined for each stage. Guidelines are also provided for management of women with heart disease during the antepartum, intrapartum and postpartum periods, including anesthesia considerations.
Adjuvant therapy, also known as adjunct therapy or add-on therapy, is therapy given in addition to the primary or initial therapy to maximize its effectiveness.
Add-ons have become ubiquitous with the process of assisted reproduction (ART) which is markedly more complex than it was at its inception.
Ovarian stimulation for assisted reproductive technology(ART) cycle aims to provide multiple pre-ovulatory follicles for oocyte collection.
The components of a conventional ART cycle-
Induction of multi-follicular growth with exogenous gonadotropins.
Prevention of endogenous leutinizing hormone (LH) surge by using Gonadotropin releasing hormone(GnRH) analogs.
inducing endogenous LH surge or mimicking it with exogenous human chorionic gonadotropin(hCG) for oocyte maturation.
This concept is known as “CONTROLLED OVARIAN STIMULATION”
“Psychiatry and the Humanities”: An Innovative Course at the University of Mo...Université de Montréal
“Psychiatry and the Humanities”: An Innovative Course at the University of Montreal Expanding the medical model to embrace the humanities. Link: https://www.psychiatrictimes.com/view/-psychiatry-and-the-humanities-an-innovative-course-at-the-university-of-montreal
Summer is a time for fun in the sun, but the heat and humidity can also wreak havoc on your skin. From itchy rashes to unwanted pigmentation, several skin conditions become more prevalent during these warmer months.
Travel vaccination in Manchester offers comprehensive immunization services for individuals planning international trips. Expert healthcare providers administer vaccines tailored to your destination, ensuring you stay protected against various diseases. Conveniently located clinics and flexible appointment options make it easy to get the necessary shots before your journey. Stay healthy and travel with confidence by getting vaccinated in Manchester. Visit us: www.nxhealthcare.co.uk
5-hydroxytryptamine or 5-HT or Serotonin is a neurotransmitter that serves a range of roles in the human body. It is sometimes referred to as the happy chemical since it promotes overall well-being and happiness.
It is mostly found in the brain, intestines, and blood platelets.
5-HT is utilised to transport messages between nerve cells, is known to be involved in smooth muscle contraction, and adds to overall well-being and pleasure, among other benefits. 5-HT regulates the body's sleep-wake cycles and internal clock by acting as a precursor to melatonin.
It is hypothesised to regulate hunger, emotions, motor, cognitive, and autonomic processes.
The Children are very vulnerable to get affected with respiratory disease.
In our country, the respiratory Disease conditions are consider as major cause for mortality and Morbidity in Child.
The biomechanics of running involves the study of the mechanical principles underlying running movements. It includes the analysis of the running gait cycle, which consists of the stance phase (foot contact to push-off) and the swing phase (foot lift-off to next contact). Key aspects include kinematics (joint angles and movements, stride length and frequency) and kinetics (forces involved in running, including ground reaction and muscle forces). Understanding these factors helps in improving running performance, optimizing technique, and preventing injuries.
Discover the benefits of homeopathic medicine for irregular periods with our guide on 5 common remedies. Learn how these natural treatments can help regulate menstrual cycles and improve overall menstrual health.
Visit Us: https://drdeepikashomeopathy.com/service/irregular-periods-treatment/
sperm assessment- traditional and novel approaches.pptx
1. SPERM ASSESSMENT – TRADITIONAL AND
NOVEL APPROACHES
Dr. Deepeka. T. S.
Fellow, Reproductive Medicine
CIMAR, KOCHI
2. Male factor infertility
Recognition of male factors has been
increasing over years.
Contributes directly or indirectly to 20-70%
of infertility in couples.
3. SEMEN ANALYSIS IS THE KEY INVESTIGATION IN
ASSESSING MALE FERTILITY
Overall
functionality of
sperm production
by testes
Secretory
activity of
accessory
glands
Patency of
genital tract
4.
5. • The latest WHO recommendations,2010 are based on
semen parameters from approximately 2000 fertile men,
from eight countries and three continents, whose
partners achieved pregnancy within 12 months of
unprotected sexual intercourse.
• Pitfalls- huge shift in the lower reference values, one
sided criteria.
• Reference limits shouldn’t be over-interpreted
• Interpret along with clinical history and physical
examination.
7. Structure of a mature spermatozoon
• 5 μm length
• Haploid
nucleus
• Acrosomal cap
• Mictochondria
-ATP
• Flagellum
8. • Gene directing spermatogenesis -Y Chromosome
• Approximately 70 days to complete
• 12-21 days- transport – testes through epididymis to
ejaculatory duct.
• 100-200 million sperms/day
• 1 trillion-during normal
reproductive life span
11. Specimen collection for diagnostic
purposes
• Collection of semen
should ideally mimic the
in vivo situation.
• Under natural conditions-
ejaculation happens in a
sequential manner.
12. • During ejaculation the first semen fractions voided
are mainly sperm-rich prostatic fluids, whereas later
fractions are dominated by seminal vesicular fluid
(Björndahl & Kvist, 2003).
• Therefore, losing the first (sperm-rich) portion of
the ejaculate has more influence on the results of
semen analysis than does losing the last portion.
– WHO, 2010.
14. Period of abstinence
• The sample should be collected after a minimum of 2
days and a maximum of 7 days of sexual abstinence.
If additional samples are required, the number of days of
sexual abstinence should be as constant as possible at
each visit.
WHO,2010.
• Ideally, 2-3 days of sexual abstinence.
Shorter abstinence period- ART procedures.
15. Collection at the laboratory
• A private room near the laboratory(limits the exposure of the
semen to fluctuations in temperature and to control the time
between collection and analysis).
• Obtained by masturbation and ejaculated into a clean, wide-
mouthed, non-toxic container made of glass or plastic.
• Without using lubricants or condoms with spermicidal agents.
• Clear instructions & documentation.
• The specimen container should be kept at ambient
temperature, between 20 °C and 37 °C.
16. Home collection
• In exceptional circumstances, such as a demonstrated inability
to produce a sample by masturbation.
• Clear instructions.
• Recording the time of semen production and deliver the
sample to the laboratory within 1 hour of collection.
• Documentation of loss of fractions/incomplete collection.
• During transport to the laboratory, the sample should be kept
between 20 °C and 37 °C.
• Inability to collection- coitus interruptus, non spermicidal
condoms(commercially available), not a reliable method.
WHO 2010.
17. Sample handling at laboratory
• Record the details- name, identification number, time
of collection, time received, date, place and method
of collection, period of abstinence, completion of
collection, time interval between collection and
examination.
• Universal precautions to avoid contamination.
• Analysis to be completed within 60 minutes.
18. Macroscopic Evaluation
• Should begin with a simple inspection soon after
liquefaction, preferably at 30 minutes, but no longer
than 1 hour after ejaculation, to prevent
dehydration or changes in temperature from
affecting semen quality.
19. Liquefaction
• Immediately after ejaculation, semen is typically a semisolid
gel like coagulated mass(cross-linking of seminal vesicle
proteins).
• Within a few minutes at room temperature, the semen usually
begins to liquefy (become thinner), becomes more
homogeneous and quite watery.
• The complete sample usually liquefies within 15 minutes at
room temperature, although rarely it may take up to 60
minutes or more.
• Continuous gentle mixing or rotation of the sample container.
20. Delayed liquefaction
• If liquefaction does not occur within 30 min, incubate for another
30 min at room temperature or in 37º c incubator.
• If semen fails to liquefy within 60 min- delayed liquefaction.
• Methods to liquefy
– addition of an equal volume of physiological medium (e.g.
Dulbecco’s phosphate-buffered saline
– repeated (6–10 times) gentle passage through a blunt gauge 18
or gauge 19 needle attached to a syringe.
– digestion by bromelain, a broad-specificity proteolytic enzyme
WHO 2010.
21. Appearance of the ejaculate
• A normal liquefied semen sample has
a homogeneous, grey-opalescent
appearance.
• It may appear less opaque if the sperm
concentration is very low.
• The colour may also be different, i.e.
red-brown when red blood cells are
present (haemospermia), or yellow in
a man with jaundice or taking certain
vitamins or drugs.
22. Semen viscosity
• Allowing the semen to drop by gravity.
• A normal sample leaves the pipette in small
discrete drops.
• If viscosity is abnormal, the drop will form a
thread more than 2 cm long.
• High viscosity – interferes with determination of
sperm motility, sperm concentration, detection
of antibody-coated spermatozoa.
• Hyperviscosity treated – physiological medium,
repeated pipetting, α-chymotrypsin and
incubate extra 10 min at 37º c
23. Semen volume
• Precise measurement of volume is essential in any
evaluation of semen, because it allows the total number
of spermatozoa and non-sperm cells in the ejaculate to
be calculated.
• Best measured by weighing the sample in the vessel in
which it is collected.
– Collect the sample in a pre-weighed, clean,
disposable container.
– Weigh the vessel with semen in it.
– Subtract the weight of the container.
– Calculate the volume from the sample weight,
assuming the density of semen to be 1 g/ml
24. • Alternatively, the volume can be measured directly.
– Collect the sample directly into a modified graduated glass measuring
cylinder with a wide mouth.
– These can be obtained commercially.
– Read the volume directly from the graduations (0.1 ml accuracy).
• Measuring volume by aspirating the sample from the specimen container
into a pipette or syringe, is not recommended, because not all the
sample will be retrieved and the volume will therefore be
underestimated.
• The volume lost can be between 0.3 and 0.9 ml
(Brazil et al., 2004a; Iwamoto et al., 2006; Cooper et al., 2007).
25. The lower reference limit for semen volume is
1.5 ml
Low semen volume
• ejaculatory duct obstruction.
• congenital bilateral absence of
the vas deferens (CBAVD)
• poorly developed seminal vesicles
• partial retrograde ejaculation
• androgen deficiency.
• loss of a fraction of the ejaculate,
short abstinence(false low)
High semen volume
• may reflect active exudation
in cases of active
inflammation of the
accessory organs.
de la Taille et al., 1998; Daudin et al., 2000; von
Eckardstein et al., 2000; Weiske et al., 2000
26. Semen pH
• The pH should be
measured after
liquefaction at a uniform
time, preferably after 30
minutes, but in any case
within 1 hour of
ejaculation since it is
influenced by the loss of
CO2 that occurs after
production.
27. WHO lower reference value- 7.2
• Higher than 8 is abnormal.
• If the pH is less than 7.0 in a semen sample with low
volume and low sperm numbers= ejaculatory duct
obstruction, CBAVD, poorly developed seminal
vesicles.
28. Initial microscopic investigation
• Gives information about quality of sperms as well as
presence of non-sperm cells.
• The liquefied sample is thoroughly mixed in the
original container and only about 50 µL is used for
evaluation.
• A phase-contrast microscope
• Involves scanning the preparation at a total
magnification of ×100.
29. Aggregation of spermatozoa
• The adherence either of immotile spermatozoa to each
other or of motile spermatozoa to mucus strands, non-
sperm cells or debris is considered to be nonspecific
aggregation.
31. • The presence of agglutination is suggestive of the
presence of anti-sperm antibodies; further testing is
required.
• Severe agglutination can affect the assessment of
sperm motility and concentration.
32. Sperm motility
• Should be assessed as soon as possible after
liquefaction of the sample, preferably at 30 minutes,
but in any case within 1 hour, following ejaculation,
to limit the deleterious effects of dehydration, pH or
changes in temperature on motility.
• phase-contrast optics at ×200 or ×400 magnification
33. Categories of sperm movement
• Progressive motility (PR): spermatozoa moving actively,
either linearly or in a large circle, regardless of speed.
• Non-progressive motility (NP): all other patterns of
motility with an absence of progression, e.g. swimming in
small circles, the flagellar force hardly displacing the
head, or when only a flagellar beat can be observed.
• Immotility (IM): no movement.
34. • The previous edition of WHO manual recommended that
progressively motile spermatozoa should be categorized
as rapid or slow, with a speed of >25 m/sec at 37 °C as
“grade a” spermatozoa.
• However, it is difficult for technicians to define the
forward progression so accurately without bias.
(Cooper & Yeung, 2006).
• When discussing sperm motility, it is important to specify
total motility (PR + NP) or progressive motility (PR)
35. • Prepare a wet preparation approximately
20 m deep
• Evaluate at least 200 spermatozoa in a total
of at least five fields in each replicate, in
order to achieve an acceptably low
sampling error.
• Assess only intact spermatozoa (defined as
having a head and a tail;
• Scan and count quickly to avoid
overestimating.
36.
37. Lower reference limit WHO, 2010.
• Total motility (PR + NP) is 40%
• Progressive motility (PR) is 32%
• The total number of progressively motile spermatozoa in the
ejaculate is of biological significance.
• This is obtained by multiplying the total number of spermatozoa in
the ejaculate by the percentage of progressively motile cells.
• Falsely low motility
– Non liquefaction
– Hyper-viscosity
– Delayed analysis.
38. Sperm concentration
• No of sperms per mL of ejaculate
• Assessed by using a hemocytometer.
• Laborious and time consuming.
• Proper mixing with phosphate buffered saline(PBS) should
be made.
• Assessing the samples within 10–15 minutes
• Counting at least 200 spermatozoa per replicate.
40. Sperm concentration, sperm count
• Sperm concentration- no of spermatozoa per ml
• Total sperm count- sperm concentration x volume
• Both are related to both time to pregnancy and
pregnancy rates and are predictors of conception
(Slama et al., 2002) (WHO,1996; Zinaman et al., 2000) (Bonde et al., 19980Larsen et al.,
2000).
When semen volume is small and fewer spermatozoa are counted than
recommended, the precision of the values obtained will be significantly
reduced.
41. • Lower reference limit- 15 × 10 6 spermatozoa per ml,
39 × 10 6 spermatozoa per ejaculate.
• Azoospermia- Absence of sperms in ejaculate in at least
2 samples on different days.
• The term azoospermia can only be used if no
spermatozoa are found in the sediment of a centrifuged
sample.
• Endocrine evaluation useful in oligo-zoospermia and
non-obstructive azoospermia.
42. Undifferentiated Round Cells
• Includes immature cells and leukocytes.
• >1 X 10 6/mL is abnormal.
• Immature germ cells indicate testicular damage.
• Leukocytospermia is suggestive of inflammation.
• Characterisation of leukocytes confirmed by
assessing peroxiadase acitivity with Endtz test.
43. Morphology
• Crucial, especially of ART.
• Complex and difficult parameter
• Abnormal morphology- abnormal spermatogenesis, sperm
maturation.
• Impact on chromatin condensation, acrosome reaction, sperm
motility.
• For morphological analysis, it is customary to prepare semen
smears that are air-dried before fixation and staining.
• Lower reference value- 4%. Teratozoospermia < 4% of
normal.
44. The concept of normal spermatozoa
• The human zona pellucida selects a subpopulation of
morphologically similar spermatozoa, but such “zona-
preferred” spermatozoa display a wider range of forms
(Liu et al., 1990; Garrett et al., 1997).
• The percentage of motile spermatozoa in semen from fathers
displaying “zona-preferred” morphology is also low (8–25%)
(Liu et al., 2003).
• The range of normal values for both fertile and infertile
men is likely to be 0–30%, with few samples exceeding
25% normal spermatozoa.
(Menkveld et al., 2001)
45.
46. Staining methods
• The use of the Papanicolaou, Shorr
or Diff- Quik stain is
recommended.
• The head is stained pale blue in the
acrosomal region and dark blue in
the post-acrosomal region.
• The midpiece may show some red
staining and the tail is stained blue
or reddish.
47. • Slides stained using the Papanicolaou procedure can
be permanently mounted and stored for future use
in internal quality control programmes.
• If stored in the dark, they should be stable for
months or years.
49. Vitality
• The evaluation of vitality differentiates live or dead
immotile sperms.
• Especially important for samples with less than about
40% progressively motile spermatozoa.
• Principle-The percentage of live spermatozoa is assessed
by identifying those with an intact cell membrane, from
dye exclusion or by hypotonic swelling.
• Lower reference limit- 58%.
51. Hypo-0smotic swelling test
• Based on the principle
that only living cells
with intact membrane
will swell under hypo-
osmotic conditions –
150 mOsmol/L
• Doesn’t kill sperms, can
be used for ART.
52. Endtz test
• Indicated when undifferentiated round cell >1 X 10 6/mL
• To detect peroxidase within neutrophils
• Ortho-toluidene stain
• A concentration of leukocytes ≥1 x 10 6/mL- positive.
• Inflammation and infective etiology, elevated anti-
sperm antibody levels, and high levels of ROS.
53. Fructose test
• Fructose in semen is a marker for seminal vesicles
function.
• Usually performed to localize the level of obstructive
azoospermia in men with low ejaculate volume.
• Lower reference limit = 13 µmol.
• Low fructose, low semen volume and low pH=
obstructive azoospermia.
• Low fructose is characteristic of partial retrograde
ejaculation.
56. Computer-Assisted Sperm
Analysis(CASA)
• Main objective- reducing the
subjectivity , possibility
decreasing the intra- and inter-
observer variability.
• Reduced the labor-intensive and
time consuming-nature of
standard techniques.
• Principle- etablishing a centroid,
evaluating cell motion based on
centroid trajectory.
• Bulk movements.
57. What semen parameters can be evaluated by
CASA?
• Sperm concentration, vitality, motility
• Very few systems – morphology- computer aided sperm
morphometric assessment(CASMA).
• Recent CASA systems have incorporated DNA
fluoroescent staining and fluorescent microscopy.
• The precision of CASA decreases outside an optimum
range of sperm concentration between 2 and 50 x
106/mL.
58. Advantages of CASA
• Decreases subjectivity
• More rapid analysis of larger sample with reduced
sampling error.
• Introduction of novel concepts- kinematics – may
reveals overall quality of sperms
• Images , recorded pictures are helpful in
standardization and quality assurance.
59. Disdvantages of CASA
• Limited number of parameters
• Doubtful cost-effectiveness
• Equipment maintenance, training of personnel
• Biological factors like hyperviscosity, sperm
agglutination hinders CASA interpretation.
• Technical errors, lack of internal quality control
• Lack of accuracy
60. Applicability of CASA
• A significant association between CASA analysis of sperm
concentration and progressive motility with fertilization
rates and time to conception.
• Only small number of studies provide the evidence.
• Many laboratories have included CASA as control for
comparison to standard technical analysis rather than a
stand-alone replacement technique.
• Many studies are needed on CASA using large samples.
61. Home test
• Commercially available
• Colour change antibody
reaction
• Positive when sperm
concentration is at lest
20 x10 6/mL
63. • Enthusiasm in male factor infertility has decreased since
the introduction of ICSI.
• Abnormal semen parameter cannot adequately predict
male fertility potential
• A significant proportion of men with apparently normal
semen parameter are classified as idiopathic male
infertility.
• The attempt to detect abnormal sperm function is one of
the major challenges to fertility experts.
64. Do they better discriminate
infertile and fertile men ?
DNA
damage
assessment
acrosome
reaction
test
Free
radical
assessme
nt
67. Methods to measure Reactive Oxygen Species
• Chemiluminescence – most common –
this method is based on production of
light through reaction between a probe
and ROS.
• Flow cytometry – non fluorescent
probes.
• Thiobarbituric Acid Reactive
Substances(TBARS)
• Nitroblue Tetrazolium (NBT)
68.
69.
70. ROS-TAC Score
• Overall level of oxidative stress by balance between
pro-oxidants and anti-oxidants.
• ROS-TAC score has higher predictive value than ROS
or TAC alone.
• Infertile men with higher ROS-TAC score are more
prone to initiate pregnancies than those with low
scores.
71. Mioxyx system – Oxidation-reduction
potential(ORP)
• ORP values higher than the
established reference values
are indicative of oxidative
stress.
• ORP valueof 2.59 mV/million
sperm/mL is a good predictor
of oligozoospermia.
• Can be use for fresh and
frozen samples.
• Results available in 2 min
72. Clinical applicability of ROS/TAC/ORP
• Elevated ROS were detected in semen of 25-40% of
infertile men.
• Some infertile men demonstrate 30-43 % decrease in
TAC.
• Increased ROS levels correlated with abnormal sperm
parameters and DNA fragmentation, leukocyte
concentration and incidence of apoptotic/necrotic
sperms.
• Elevated ROS – increased time to natural conception,
impaired IVF pregnancy rates, recurrent miscarriages.
73. • Oral antioxidant therapy, varicocelecotmy and life style
modifications can be attempted and response assessed.
• Modifications of media by applying antioxidants to reduce OS.
• Analysing ROS and TAC can be complementary to conventional
semen analysis in ART treatment and oral antioxidant can be
used before ART.
• ORP with its high sensitivity, specificity, accuracy and low
intra-inter observer variability has the potential in
distinguishing infertile and fertile men, additional research is
needed.
74. Sperm DNA damage
• Sperm DNA integrity is vital for fertilization and
trasmission of genetic material to the offspring.
• The DNA of mammalian sperm is the mostly known
compact eukaryotic DNA(packaged six times more tightly
than the somatic cells).
• DNA damage can occur not only during spermatogenesis,
but epididymal transit and collection for ART.
• Single stranded breaks and double stranded breaks are
commonly termed DNA fragmentation.
75.
76. Methods to measure sperm DNA damage
1. Terminal deoxynucleotidyl transferase-mediated
dUTPnick end labeling assay (TUNEL)
– based dUTP binding to the 3-OH break ends of single
and double stranded DNA breaks.
– Fluorescent signal = percent of DNA fragmentation.
– cut-off values- 17 %
– High specificity (92%), high PPV(91%)
– Available as commercial kit
77. 2. Comet assay
embedding the certain number of sperm into the agar and lysing their cellular
proteins and membranes using detergent and high density salt solutions
exposed to electrophoresis
DNA fragments move towards postively charged anode.
evaluation of the comet image by fluorescence microscope or cytometer
78. • Length of comet tail and degree of fluorescence
• Prediction of fertilization rates with 93.3% specificity.
• DNA fragmentation >40 % reflects a 9.5 times higher
risk of poor fertilization.
T.R.Dias et al
79. 3. Sperm Chromatic Dispersion(SCD)
Single-cell agarose gel
electrophoresis
Acid/alkaline denaturation
Bright field or fluorescent
microscope
81. Sperm chromatin structure assay
• Principle DNA damaged sperm are more vulnerable to
heat or acid denaturation compared to the intact sperm.
• The denatured DNA damaged sperm are stained red
while the intact sperm turn into green when exposed to
acid and acridine orange stain.
• The stained sperm are analyzed by flow cytometry.
• The greatest advantage -large number of cells could be
evaluated
82. • DFI ≥30 % - high risk of low
blastocyst rates, failure to initiate
ongoing pregnancy
• DFI ≥15 % - lower fertilization
rates, ICSI recommended.
T.R.Dias et al
83. Clinical value of sperm DNA damage evaluation
• High rates of DNA fragmentaiton were correlated with decreased
semen quality.
• Sperm DNA damage is significantly increased in idiopathic and male
factor infertility.
• Sperm DNA fragmentation is associated with longer time for
natural conception, idiopathic infertility, recurrent IUI and IVF
failures and spontaneous miscarriage.
• Clinical utility proposed –
– Infertile men with normal semen analysis
– Recurrent spontaneous abortion
– To determine most suitable ART
85. Acrosome reaction test
• Invitro acrosome reaction
dysfuction can be assessed.
• Artificial acrosome reaction
induced by calcium inophore A
23187.
• Fluorescent labelled lectins and
monoclonal antibodies-used to
asses the acrosomal
membranes and contents.
• 5-30 % reacted sperms means
higher fertility potential.
T.R.Dias et al
86. Sperm mitochondrial activity
• A fluorescent probe is used to distinguish between
sperm with poorly and highly functional
mitochondria using flow cytometry.
• Only method available for assessment of sperm
mitrochondria.
• A recent study- DNA FRAG testing + sperm
mitochondrial membrane potential could be stronger
predictors of natural conception.
87. Antisperm antibodies(ASA)
• Among all male infertility cases, 10 % are associated with ASA.
• Indication – agglutination in conventional SA.
• Mixed antiglobulin reaction(MAR)
• Immunobead test(IBT)
• Clinically significant when ≥50 % are coated , when sperms are
unable to penetrate pre-ovulatory cervical mucus or sperms
demonstrated impaired fertilizing capacity.
• Controversy about predicting ART outcome.
88. Future perspectives
• Digital holography microscopy- morphology and
motility characterisation, using three-dimensional
images to delineate normal and abnormal sperm.
• Study of semen using genomics, proteomics,
metabolomics -new insight into biochemical basis of
defective semen, could be a new tool for diagnostic
and therapeutic purposes.
89. Concluding remarks
• A properly conducted standardized semen analysis
technique is of utmost importance for diagnosis and
evaluation of infertile men.
• Introduction of CASA aims to overcome limitations of
conventional SA, however, with questionable efficiency,
lack of standard guidelines, inconsistency between
different equipments and high cost.
• The necessity to over come the limitations of routine
semen analysis is quite clear.
90. • Sperm function test can be valuable complement to
conventional semen analysis to evaluate fertility,
counsel the couple as well as predict reproductive
outcome by natural conception or ART.
• Sperm DNA assessment may have role in idiopathic
infertility, recurrent IUI and IVF failures and
spontaneous miscarriage in choosing the best
method of ART.
91.
92. References
• WHO laboratory manual for the Examination and processing
of human semen(FIFTH EDITION)
• Dias T.R., Cho CL., Agarwal A. (2019) Sperm Assessment:
Traditional Approaches and Their Indicative Value. In: Nagy Z.,
Varghese A., Agarwal A. (eds) In Vitro Fertilization. Springer,
Cham. https://doi.org/10.1007/978-3-319-43011-9_22
• Dias T.R., Cho CL., Agarwal A. (2019) Sperm Assessment: Novel
Approaches and Their Indicative Value. In: Nagy Z., Varghese
A., Agarwal A. (eds) In Vitro Fertilization. Springer, Cham.
https://doi.org/10.1007/978-3-319-43011-9_23