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SCSA Diagnostics es la pionera en el análisis del ADN espermático, utilizando investigaciones establecidas y estudios clínicos en curso durante más de 25 años para realizar una de las pruebas de fragmentación del ADN espermático más avanzadas disponibles.
El desarrollo de la prueba SCSA se presentó en la portada de la reconocida revista Science en diciembre de 1980.
Es importante saber que hay algunos laboratorios que ofrecen una prueba similar a SCSA. Sin embargo, no se trata de la precisión de la prueba SCSA®
La fragmentación del ADNdel esperma es una de las causas paternas más importantes de fracasos reproductivos repetidos. Alrededor del 20-30% de los hombres que visitan una clínica de fertilidad tienen un nivel de fragmentación del ADN espermático que puede conducir a la falla del embarazo.
Spano et al. 2000 La línea discontinua muestra el umbral clínico para la fragmentación del ADN espermático y las posibilidades de embarazo.
Algunos autores han publicado que las muestras de semen que contienen más del 27-30% de espermatozoides con fragmentación del ADN (% del Índice de Fragmentación del ADN) tienen entre 7 y 10 veces menos probabilidades de lograr el embarazo mediante relaciones sexuales o IIU, y alrededor de 1,5-2,0 veces menos probabilidades mediante FIV/ICSI.
Giwercman A et al. (2009 en imprenta, Int. J. Androl) incluyeron 137 hombres de parejas con infertilidad comprobada sin factores maternos conocidos y 137 hombres con fertilidad comprobada.
En hombres con parámetros de semen normales (concentración, movilidad y morfología del espermatozoide), el riesgo de infertilidad aumentó 5 veces con valores de Índice de fragmentación del ADN superioresal 20%.
Sin embargo, si alguno de los parámetros de semen era anormal, el riesgo de infertilidad se incrementó 16 veces incluso con valores de Índice de fragmentación de ADN superiores al 10%.
Los autores concluyeron que "el Índice de fragmentación del ADN es un factor predictor de la probabilidad de concepción natural, incluso cuando los parámetros seminales clásicos se encuentran dentro del rango normal. El análisis SCSA complementa la planificación del tratamiento para parejas con infertilidad comprobada"
La integridad del ADN espermático es continuamente desafiada por factores endógenos y exógenos, aunque varios mecanismos para reparar y proteger el ADN del daño están activos en las células humanas (Hoeijmakers, 2009). En particular, la preservación de la integridad del ADN tiene un papel importante en la transmisión del genoma a la próxima generación. En estas células, se inducen fisiológicamente rupturas de doble cadena del ADN durante la espermatogénesis y espermiogénesis para facilitar el entrecruzamiento meiótico y la sustitución de histonas-protaminas, respectivamente (Rathke et al., 2014). Además de este daño "fisiológico" del ADN, otros factores exógenos y endógenos pueden afectar la integridad del ADN durante la maduración/almacenamiento de los espermatozoides en el epidídimo (Moustafa et al., 2004; Ramos et al., 2004; Sakkas et al., 2002). Estudiar la integridad del ADN podría ser un paso fundamental en la evaluación de la competencia funcional del espermatozoide (Lewis et al., 2008). Por lo tanto, el análisis del daño del ADN espermático podría ser crucial tanto para el diagnóstico de la infertilidad como para predecir el éxito de la tecnología de reproducción asistida (ART, por sus siglas en inglés). En el contexto de la FIV, esta evaluación es de particular importancia ya que evita las barreras de selección natural de la concepción, aumentando la probabilidad de que los espermatozoides con un daño significativo del ADN transmitan las anomalías genéticas al recién nacido (Host et al., 2000). Por lo tanto, varios estudios han evaluado el papel predictivo del daño del ADN espermático para el resultado de la tecnología de reproducción asistida o la selección de espermatozoides. El daño en el ADN del espermatozoide puede ser resultado de cinco mecanismos patogénicos diferentes (Perrin et al., 2011; Sakkas y Alvarez, 2010).
Índice de Fragmentación del ADN :
El Índice de Fragmentación del ADN es la medida de la fragmentación del ADN espermático y un indicador de la calidad del esperma.
Células con Alta Tinción del ADN: Porcentaje de espermatozoides con proteínas nucleares y estructuras de cromatina anormales (espermatozoides inmaduros).
La prueba SCSA® se realiza utilizando un dispositivo láser llamado citómetro de flujo que mide 250 espermatozoides por segundo. El resultado de la prueba se basa en la medición de aproximadamente 10.000 espermatozoides.
PRUEBA DE CRIBADO : Especialmente indicada para parejas con un ansiado deseo de tener hijos
INTENTOS INFRUCTUOSOS DE IIU/FIV/ICSI
ABORTOS ESPONTÁNEOS RECURRENTES
BAJA CALIDAD Y DESARROLLO EMBRIONARIO
INFERTILIDAD INEXPLICABLE: Parejas que desean tener hijos y han obtenido pruebas diagnósticas sin resultados significativos
HOMBRES PROPENSOS A UN ALTO ESTRÉS OXIDATIVO:
ESTILO DE VIDA: tabaquismo, consumo de drogas, consumo excesivo de alcohol, obesidad, estrés
NUTRICIÓN: Aunque muchas veces ignorada, hemos observado mejoras drásticas en la fragmentación del ADN espermático después de un análisis detallado y la corrección de la nutrición diaria.
EXPOSICIÓN AL CALOR: por ejemplo, baños de hidromasaje, trabajar con un portátil en el regazo, fiebre, etc.
EXPOSICIONES OCUPACIONALES: productos químicos, pesticidas, fertilizantes, tintes, etc.
DIABETES: Factor de riesgo independiente para el aumento de la fragmentación del ADN espermático.
MEDICAMENTOS: Incluso los medicamentos más inofensivos a veces han demostrado causar fragmentación del ADN espermático.
TOXINAS AMBIENTALES / Contaminación del aire: Aunque pueda parecer general, se debe realizar un examen atento a la exposición.
VARICOCELE: Una vez diagnosticado ya sea por examen o ultrasonido, la prueba SCSA puede ayudar en la decisión si la cirugía será útil.
CÁNCER Y TRATAMIENTOS CONTRA EL CÁNCER (quimioterapia y radioterapia): causan daño directo al ADN de las células.
ENFERMEDADES CRÓNICAS: a menudo causan un aumento del estrés oxidativo que lleva a un aumento del daño al ADN.
ENFERMEDADES ANDROLÓGICAS / Infecciones del tracto urogenital y leucocitospermia: la infección / inflamación local causa daño directo al ADN del espermatozoide a través de la liberación de especies reactivas de oxígeno (ROSpor sus siglas en inglés).
EDAD > 40 AÑOS: Se ha estudiado ampliamente que cuanto mayor es el hombre, mayor es la fragmentación del ADN.
LUBRICANTES: ccausan daño directo al ADN del espermatozoide.
La prueba SCSA® es realizada utilizando un dispositivo láser llamado citómetro de flujo. Básicamente, el citómetro de flujo mide 250 espermatozoides por segundo con alta precisión y proporciona un resultado estadísticamente sólido basado en datos de 2x 5000 espermatozoides por muestra. La precisión del citómetro de flujo combinada con la bioquímica precisa de la prueba SCSA permite mediciones altamente precisas de la integridad del ADN y de la cromatina del espermatozoide.
Las células se tiñen con un tinte fluorescente de ADN (naranja de acridina) y se fuerzan a través de un canal de vidrio en una suspensión líquida. Cuando las células pasan por el rayo láser, la luz láser hace que el tinte emita luz fluorescente de un color específico..
En la prueba SCSA®, debido a la naturaleza única del tinte naranja de acridina, los espermatozoides que emiten fluorescencia verde tienen niveles indetectables de ADN fragmentado, mientras que los espermatozoides que emiten fluorescencia de amarilla a roja tienen niveles moderados a altos de ADN fragmentado.
La fertilidad masculina se examina tradicionalmente mediante pruebas de esperma (espermiogramas) que registran la concentración, la motilidad y la morfología de los espermatozoides. Sin embargo, numerosos estudios en el último medio siglo demuestran claramente que estos parámetros no predicen el embarazo, ya que los hombres fértiles e infértiles tienen valores superpuestos (con la excepción de la azoospermia). Los espermiogramas clásicos son mediciones microscópicas de factores externos de los espermatozoides, mientras que la prueba SCSA mide factores internos del núcleo que no se pueden medir utilizando el microscopio óptico.
La prueba SCSA es una prueba muy precisa que requiere un citómetro de flujo que es inaccesible para la mayoría de las clínicas de fertilidad debido a los altos costos de capital y personal capacitado. Por lo tanto, las muestras de semen recolectadas por usted o en el hogar del paciente pueden ser criopreservadas en su estado natural.
Puede informarnos por correo electrónico acerca del transporte deseado. Le enviaremos un contenedor preenfriado; las muestras de semen son almacenadas y enviadas de regreso a nuestro laboratorio.
El informe del SCSA estará disponible para usted a través de un sitio web seguro aproximadamente 4 días laborales después de que la muestra llegue a nuestro laboratorio.
Las muestras deben obtenerse después de un período de abstinencia de 24 a 48 horas, mediante masturbación en recipientes estériles de plástico para las muestras. Después de la licuefacción de la muestra, la concentración, la motilidad y la morfología del esperma son determinadas. Anote los parámetros en la carta de porte.
La concentración del esperma no debe ser inferior a 500,000/ml . De lo contrario, pueden producirse resultados inconclusos.
La criopreservación debe realizarse en un plazo máximo de 1 hora después de la entrega, ya que el Índice de fragmentación del ADN puede alterarse después de un período de espera más largo.
Criopreservación de las muestras:
Las muestras deben ser congeladas lo antes posible después de la licuefacción para minimizar la exposición al aire, que puede causar daños por especies reactivas de oxígeno.
Prepare 2 crioviales (tubos de 1-2 ml): Etiquételos con el nombre del paciente y la fecha de recolección de la muestra.
Congelación rápida:
Pipetee 0,25 ml o más del eyaculado nativo licuado en cada tubo criogénico.
¡No se requieren crioprotectores!
Coloque los tubos bien cerrados en posición vertical en el soporte de metal suministrado y coloque esto directamente en el recipiente criogénico preenfriado. El contenedor debe cerrarse rápidamente.
El contenedor se cierra con la abrazadera suministrada y se coloca en el recipiente de plástico, al que se adjunta la carta de porte para que sea claramente visible.
Nota: si la concentración de espermatozoides es suficiente, menos de 0,5 ml (dos alícuotas de 0,25 ml) de eyaculado puede ser suficiente para la prueba SCSA®. Para realizar la prueba, necesitamos dos alícuotas de 0,20 ml con una concentración > 0,5 m/ml.
Por favor, póngase en contacto con nuestro laboratorio si tiene alguna pregunta sobre el volumen requerido para una muestra de una concentración específica.
Envío de las muestras:
Enviamos un contenedorpreenfriado. Los contenedores almacenan nitrógeno líquido en una especie de "esponja" dentro del recipiente y así garantizan el transporte seguro de sus muestras criopreservadas en nitrógeno en fase gaseosa. La baja temperatura en el contenedor se puede mantener durante 7 días. Por lo tanto, es importante enviarlo de vuelta lo antes posible utilizando la etiqueta de envío proporcionada.
En el espacio entre el contenedor y la caja de envío hay un sobre con una abrazadera y un soporte para ampollas. También contiene una etiqueta de envío y una funda correspondiente quese debe atar al recipiente para el envío.
El interior del contenedor es muy frío (-196°C). Para evitar la congelación, debe usar guantes al tocar cualquier cosa que esté dentro del contenedor.
La prueba SCSA divide a los espermatozoides en cinco poblaciones:
El porcentaje de espermatozoides con cromatina inmadura. Los espermatozoides con Células con Alta Tinción del ADN presentan menos condensación de la cromatina, lo que resulta en una mayor tinción del ADN. Algunos estudios han demostrado que, si el porcentaje deCélulas con Alta Tinción del ADN es superior al 36%, no se ha producido ningún embarazo (Menezo).
Los valores del Índice de fragmentación del ADN se dividen en cuatro categorías estadísticas y pueden proporcionar las siguientes previsiones:
Índice de fragmentación del ADN<15% = resultados de embarazo muy buenos a buenos
Índice de fragmentación del ADN de > 15% a <25% = resultados de embarazo buenos a moderados
Índice de fragmentación del ADN de > 25% a <40% = resultados de embarazo moderados a malos
Índice de fragmentación del ADN40% = resultados muy malos de embarazo Todos los datos deben ser interpretados sin la influencia de factores maternos.
Además, el hallazgo muestra una representación gráfica del porcentaje del Índice de
Fragmentación del ADN y del porcentaje de Células con Alta Tinción del ADN.
El gráfico muestra que el 76% de los espermatozoides medidos no presentan fragmentación del ADN y el 24% restante presenta una fragmentación moderada-alta del ADN (Índice de fragmentación del ADN = 24%). Es posible concebir de forma natural o con la ayuda de una IIU.
Los datos actuales muestran que la probabilidad de un embarazo exitoso es significativamente menor cuando la proporción de espermatozoides con ADN fragmentado es > 25%. Sin embargo, un Índice de fragmentación del ADN> 25% no descarta el embarazo. El umbral de 25% es un umbral estadístico. Si un hombre tiene un Índice de fragmentación del ADNconstante >25%, se le incluye en un grupo estadístico que ha demostrado en ensayos clínicos que tarda más tiempo en lograr el embarazo natural y requiere más ciclos de FIV. Se asocia una tasa aumentada de abortos espontáneos con esto o el embarazo no se produce en absoluto.
Evaluación de la calidad del ADN espermático
Si el Índice de fragmentación del ADN ylasCélulas con Alta Tinción del ADN se encuentran dentro del rango normal, es posible el embarazo de manera natural o mediante inseminación (ignorando los factores de la pareja que influyen en la fertilidad). Esto significa que el ADN espermático muestra una alta estabilidad.
El Índice de fragmentación del ADN ha aumentado significativamente. Esto indica que el ADN espermático es muy frágil y tiende a la fragmentación del ADN.
La estabilización del ADN puede lograrse mediante:
En algunos casos, puede ser necesario cambiar de IIU a FIV/ICSI para aumentar las posibilidades de un resultado positivo en el embarazo.
The histogram shows the DFI (DNA Fragmentation Index) measured in a semen sample.
DFI scores are classified into four statistical categories and can predict the following (ignoring partner factors affecting fertility):
< 15% DFI = Very good to good pregnancy outcomes
> 15% to < 25% DFI = Good to moderate pregnancy outcomes
> > 25% to < 40% DFI = Moderate to poor pregnancy outcomes
> 40% DFI = Very bad outcome of pregnancy
If the %DFI is above 25%, the current literature recommends:
Se cree que el óvulo es capaz de reparar ciertos niveles de daño en el ADN. Los niveles moderados a altos de fragmentación del ADN (Índice de fragmentación del ADN) encontrados en los espermatozoides con fluorescencia roja probablemente superan la capacidad de reparación del ADN del óvulo.
Marcos Mesequeret al. encontraron que, después de la recuperación de los óvulos, cada aumento del 10% en el Índice de fragmentación del ADN aumentó las probabilidades de no quedar embarazada en un factor de 1,31. En comparación, este efecto no se observó al utilizar óvulos de donantes, lo que indica una mayor capacidad de reparación con respecto a la fragmentación del ADN de los óvulos jóvenes.
Si el Índice de fragmentación del ADN es alto, se recomiend:
Si existe un factor de riesgo, se recomienda repetir la prueba de SCSA 3 meses después de la eliminación o corrección del factor de riesgo para evaluar su impacto.
Ejemplos:
Abstinencia de nicotina (84 días después):
Cirugía de varicocele (3 meses después):
Cese de la exposición a pesticidas (3 meses después):
El costo de la prueba es de €295. Actualmente no está cubierta por planes de salud.
El envío de sus muestras a nuestro laboratorio es gratuito.
Para valores de Índice de fragmentación del ADN >30%, le ofrecemos una segunda prueba de control gratuita, que normalmente debe realizarse 3 meses después de la primera prueba
Fragmentation of sperm DNA is one of the most important paternal causes of repeated reproductive failure. About 20-30% of men who attend a fertility clinic have a level of sperm DNA fragmentation that can lead to pregnancy failure.
Spano et al. 2000 The dashed line shows the clinical threshold for sperm DNA fragmentation and pregnancy chances
Some authors have published that semen samples containing more than 27-30% sperm with DNA fragmentations (DNA Fragmentation Index, %DFI) are 7-10 times less likely to get pregnant through intercourse or IUI, and one around 1.5 -2.0 times lower chance through IVF/ICSI
Giwercman A et al. (2009 in press, Int. J. Androl) included 137 men from couples with proven infertility with no known maternal factors and 137 men with proven fertility
In men with normal semen parameters (sperm concentration, motility and morphology), the risk of infertility was increased 5-fold with DFI values above 20%.
However, if any of the semen parameters were abnormal, the risk of infertility was increased 16-fold even with DFI values above 10%
The authors concluded that “DFI is a predictor of the likelihood of natural conception, even when classic semen parameters are within the normal range. The SCSA analysis complements treatment planning for couples with proven infertility.”
Sperm DNA integrity is continuously challenged by endogenous and exogenous factors, although different mechanisms of repairing and protecting against this damage are active in human cells (Hoeijmakers, 2009). This is particularly relevant in germ cells, which have to preserve DNA integrity to pass the genome to the next generation. In these cells, DNA double-strand breaks are physiologically induced during spermatogenesis and spermiogenesis to facilitate meiotic crossover and histone– protamine substitution, respectively (Rathke et al., 2014). Apart from this first ‘physiological’ DNA damage, other exogenous and endogenous factors could affect DNA integrity during sperm maturation and storage in the epididymis (Moustafa et al., 2004; Ramos et al., 2004; Sakkas et al., 2002). Thus, DNA integrity is constantly at risk and its assessment could be a fundamental step in the evaluation of sperm functional competence (Lewis et al., 2008). Hence, sperm DNA damage evaluation could be crucial for both infertility diagnosis and prediction of ART success. In the setting of IVF this evaluation plays a peculiar role because the natural selection barriers of conception are bypassed, increasing the possibility of spermatozoa with significant DNA damage transmitting the genetic aberrations to the newborn (Host et al., 2000). Thus, several trials have evaluated the predictive role of sperm DNA damage for either ART outcome or sperm selection.
DFI (DNA Fragmentation Index) :
Percentage of sperm with DNA fragmentation
HDS (High DNA Stainability Cells): Percentage of sperm with abnormal core proteins and chromatin structures (immature sperm)
The SCSA® test is performed using a laser-based device called a flow-cytometer which measures 250 sperm/second. The test result is based on the measurment of around 10,000 sperm.
LIFESTYLE: smoking, drug use, increased alcohol consumption, obesity, stress
NUTRITION: Although commonly overlooked, we have often seen drastic improvement in sperm DNA fragmentation after a detailed analysis and correction of daily nutrition.
HEAT EXPOSURE: e.g. from whirlpool baths, working with a laptop on the lap, fever, etc.
OCCUPATIONAL EXPOSURES: chemicals, pesticides, fertilizers, dyes, etc.
DIABETES: An independent risk factor for increased sperm DNA fragmentation
MEDICATION: Even the most unassuming medications have sometimes seen to cause sperm DNA fragmentation
ENVIRONMENTAL TOXINS /air pollution: Although it may appear general, a keen look into exposure should be ruled out.
VARICOCELE: Once diagnosed either by examination or ultrasound, SCSA test can help in the decision if surgery will help.
CANCER AND CANCER TREATMENTS (chemotherapy and radiotherapy) : causes direct damage to the DNA of cells.
CHRONIC DISEASES: often cause increased oxidative stress leading to increased DNA damage
ANDROLOGICAL DISEASES / Infections of the urogenital tract and leukocytospermia: Local infection / inflammation causes direct damage to the DNA in sperm through release of ROS (Reactive Oxygen Species).
AGE >40 YEARS: It has been extensively studied that the older the man, the higher the DNA fragmentation
LUBRICANTS: cause direct damage to the sperm DNA
Sperm DNA damage can result from five different pathogenic mechanisms (Perrin et al., 2011; Sakkas and Alvarez, 2010).
The SCSA® test is performed using a laser-based device called a flow cytometer. Essentially, the flow cytometer measures 250 sperm/s with high precision, providing a statistically robust result based on data from 2x 5000 sperm per sample. The precision of flow cytometry coupled with the precise biochemistry of the SCSA test provides highly accurate measurements of sperm chromatin and DNA integrity.
The cells are stained with a fluorescent DNA stain (acridine orange) and then forced through a glass channel in a liquid suspension. When the cells pass the laser beam, the laser light causes the dye to emit fluorescent light of a specific color.
In the SCSA® test, due to the unique nature of the Acridine Orange dye, sperm that emit green fluorescence have undetectable levels of fragmented DNA, while sperm that emit yellow to red florescence have moderate to high levels of fragmented DNA.
The SCSA test is a two-fold simultaneous flow cytometry measure of
in thousands of sperm in a fresh or frozen/thawed semen sample.
The extent of DNA strand breaks and abnormality of chromatin structure are related to male factor fertility, including time to couple pregnancy, IVF embryo quality, miscarriage, or infertility.
Male fertility is classically addressed by semen tests that include sperm density, motility and morphology. However, numerous studies over the past half century clearly show that, except for absence of sperm, these parameters do not predict pregnancy since fertile and infertile men have overlapping values. The classical semen tests are light microscope measures of external sperm factors while the SCSA test are measures of the internal nuclear factors not visible by light
microscopy.
The SCSA test is a high precision test that requires a flow cytometer which due to the need of a high capital expense and a trained technician is not amenable to most infertility clinics. Thus, semen samples collected, as per detailed instructions, in a clinic or in a patients’ home, can be flash frozen in a LN 2 dry shipper tank.(See support protocol 1).
Clinics can then send us an email informing us. A pre-cooled LN 2 shipper is then sent to the clinic and the semen samples are flash frozen and the LN 2 shipper then returned to a SCSA diagnostic testing lab.
Some clinics send frozen semen samples in well insulated dry ice containers.
SCSA clinical data are then sent via a secure WEB site to the physician or patient that ordered the test.
There are five main populations of sperm identified by the SCSA® test:
the percentage of sperm with immature chromatin. HDS sperm exhibit less chromatin condensation, resulting in increased DNA stainability. Some studies have shown that if the %HDS is above 36%, no pregnancies have occurred (Menezo)
Our SCSA clinical report lists four statistical categories of fertility potential. These were derived from a comprehensive study of male fertility potential without medical intervention and have been confirmed in additional studies.
< 15% DFI = Excellent to good pregnancy outcomes (excluding female infertility factors)
> 15% to < 25% DFI = good to fair pregnancy outcomes
> 25% to < 40% DFI = fair to poor pregnancy outcomes
> 40% DFI = Very poor pregnancy outcomes
Furthermore, the finding shows a graphical representation of the %DFI (DNA Fragmentation Index) and the %HDS (high DNA stainability)
The graph shows that 76% of the measured sperm shows no DNA fragmentation and 24% shows medium-high DNA fragmentation (DFI= 24%). Pregnancy naturally or with the help of an IUI is possible.
Current data shows that the probability of a successful pregnancy is significantly lower when the proportion of sperm with fragmented DNA is > 25%. Nevertheless, a DFI > 25% does not rule out a normal pregnancy. The 25% threshold is a statistical threshold. Therefore, if a man has a constant DFI >25%, he will be placed in a statistical group of men who have been shown in clinical trials to take longer to achieve natural pregnancy and more IVF cycles. An increased rate of spontaneous abortions is associated with this or pregnancy does not occur at all.
The ovum is believed to be able to repair a certain amount of DNA damage.
Moderate to high levels of DNA fragmentation (DFI) found in the red fluorescent sperm likely exceed the egg’s DNA repair capacity.
Marcos Meseguer et al found that when oocytes from patients were taken, for every 10% increase in DFI, there was a 1.31 times increase in probability of not achieving pregnancy. Whereas, the same was not seen when donor oocytes were employed suggesting the higher capability of young oocytes to repair sperm DNA damage.
REPEATED IUI / IVF / ICSI FAILURE : The reason behind the unsuccessful attempts at IUI/IVF/ICSI can be identified and managed
REPEATED MISCARRAGE : An explanation for repeated spontaneous abortions (RPL, repeated miscarriage) could be found and guided
POOR EMBRYO DEVELOPMENT AND QUALITY : The reason behind poor embryo quality could be the fragmented sperm DNA which could be corrected before the next treatment
MEN AT RISK FOR HIGHER OXIDATIVE STRESS:INITIAL SCREENING TEST : Especially for couples who have been attempting pregnancy for a long time to identify the best management strategy
UNEXPLAINED INFERTILITY : Couples desiring children with all normal diagnostic tests
In cases of high DFI, we suggest:
When a risk factor is identified, after avoidance / correction of the risk factor a repeat SCSA test after 3 months is advised.
Yes. We offer a SCSA test at half the cost for controlling the effect of avoidance / correction of a risk factor.
Please find a few examples below:
After quitting smoking for 84 days
3 months after varicocele correction surgery :
3 months after stopping pesticide exposure:
After 24-48 hours of abstinence, the samples should be taken by masturbation into sterile plastic sample containers. After liquefaction of the sample, the sperm concentration, motility and morphology are determined.
Cryopreservation should take place no more than 1 hour after delivery, since the DFI can be falsely raised after a longer waiting period.
Flash-freezing: Freeze 0.25mL or more of liquefied native ejaculate in 1-2mL cryovials. Cryoprotectants is not required! (Please freeze 2 cryovials per patient)
Samples should be frozen as soon as possible after liquefaction to minimize exposure to air, which can cause damage caused by reactive oxygen species.Cryovials should be tightly capped, labeled with two identifiers (e.g. name and date of birth) and placed in an upright position directly into the pre-chilled cryo-container. The container must be closed quickly.
Note: Cryoprotectants are not needed since quick-frozen sperm give the clearest SCSA® data. This feature is unique to mammalian sperm cells due to the highly condensed, crystalline nature of the nuclei.
Note: If the sperm concentration is adequate, less than 0.5 mL (two 0.25 mL aliquots) of semen may be sufficient for the SCSA® test. To perform the test, we require two 0.20 mL aliquots with a concentration of >0.5 million/ml. Please contact our laboratory if you have questions pertaining to the volume necessary for a sample of a certain concentration.
We ship a pre-chilled dry shipper. Dry shippers store the liquid nitrogen in porous material and thus ensure the safe transport of your cryopreserved samples in the nitrogen gas phase.
The low temperature in the container can be maintained for 7 days. It is therefore important to send it back as soon as possible using the mailed/supplied shipping label.
Note: liquid nitrogen is adsorbed by special materials inside the shipping container. Even if they are turned upside down during transport, no liquid nitrogen will leak out. Dry shippers are not considered dangerous.
After receiving the sample(s) in our laboratory, we need 4 working days to create a report, which we will send to you.
The cost of the test is 295 Euro.
* Samples are shipped in a pre-cooled shipper that we can provide either at ambient or pre-chilled temperatures. The pre-cooled shipper will be sent to the centre where you have provided your sample for the centre to ship it to us.
The test at present is not covered by the insurance.
Please write us an email to contact@scsalab.com and after contacting you, we will send toy a dry shipper for the patient sample to be sent to us.
Fragmentation of sperm DNA is one of the most important paternal causes of repeated reproductive failure. About 20-30% of men who attend a fertility clinic have a level of sperm DNA fragmentation that can lead to pregnancy failure.
Spano et al. 2000 The dashed line shows the clinical threshold for sperm DNA fragmentation and pregnancy chances
Some authors have published that semen samples containing more than 27-30% sperm with DNA fragmentations (DNA Fragmentation Index, %DFI) are 7-10 times less likely to get pregnant through intercourse or IUI, and one around 1.5 -2.0 times lower chance through IVF/ICSI
Giwercman A et al. (2009 in press, Int. J. Androl) included 137 men from couples with proven infertility with no known maternal factors and 137 men with proven fertility
In men with normal semen parameters (sperm concentration, motility and morphology), the risk of infertility was increased 5-fold with DFI values above 20%.
However, if any of the semen parameters were abnormal, the risk of infertility was increased 16-fold even with DFI values above 10%
The authors concluded that “DFI is a predictor of the likelihood of natural conception, even when classic semen parameters are within the normal range. The SCSA analysis complements treatment planning for couples with proven infertility.”
Sperm DNA integrity is continuously challenged by endogenous and exogenous factors, although different mechanisms of repairing and protecting against this damage are active in human cells (Hoeijmakers, 2009). This is particularly relevant in germ cells, which have to preserve DNA integrity to pass the genome to the next generation. In these cells, DNA double-strand breaks are physiologically induced during spermatogenesis and spermiogenesis to facilitate meiotic crossover and histone– protamine substitution, respectively (Rathke et al., 2014). Apart from this first ‘physiological’ DNA damage, other exogenous and endogenous factors could affect DNA integrity during sperm maturation and storage in the epididymis (Moustafa et al., 2004; Ramos et al., 2004; Sakkas et al., 2002). Thus, DNA integrity is constantly at risk and its assessment could be a fundamental step in the evaluation of sperm functional competence (Lewis et al., 2008). Hence, sperm DNA damage evaluation could be crucial for both infertility diagnosis and prediction of ART success. In the setting of IVF this evaluation plays a peculiar role because the natural selection barriers of conception are bypassed, increasing the possibility of spermatozoa with significant DNA damage transmitting the genetic aberrations to the newborn (Host et al., 2000). Thus, several trials have evaluated the predictive role of sperm DNA damage for either ART outcome or sperm selection.
DFI (DNA Fragmentation Index) :
Percentage of sperm with DNA fragmentation
HDS (High DNA Stainability Cells): Percentage of sperm with abnormal core proteins and chromatin structures (immature sperm)
The SCSA® test is performed using a laser-based device called a flow-cytometer which measures 250 sperm/second. The test result is based on the measurment of around 10,000 sperm.
LIFESTYLE: smoking, drug use, increased alcohol consumption, obesity, stress
NUTRITION: Although commonly overlooked, we have often seen drastic improvement in sperm DNA fragmentation after a detailed analysis and correction of daily nutrition.
HEAT EXPOSURE: e.g. from whirlpool baths, working with a laptop on the lap, fever, etc.
OCCUPATIONAL EXPOSURES: chemicals, pesticides, fertilizers, dyes, etc.
DIABETES: An independent risk factor for increased sperm DNA fragmentation
MEDICATION: Even the most unassuming medications have sometimes seen to cause sperm DNA fragmentation
ENVIRONMENTAL TOXINS /air pollution: Although it may appear general, a keen look into exposure should be ruled out.
VARICOCELE: Once diagnosed either by examination or ultrasound, SCSA test can help in the decision if surgery will help.
CANCER AND CANCER TREATMENTS (chemotherapy and radiotherapy) : causes direct damage to the DNA of cells.
CHRONIC DISEASES: often cause increased oxidative stress leading to increased DNA damage
ANDROLOGICAL DISEASES / Infections of the urogenital tract and leukocytospermia: Local infection / inflammation causes direct damage to the DNA in sperm through release of ROS (Reactive Oxygen Species).
AGE >40 YEARS: It has been extensively studied that the older the man, the higher the DNA fragmentation
LUBRICANTS: cause direct damage to the sperm DNA
Sperm DNA damage can result from five different pathogenic mechanisms (Perrin et al., 2011; Sakkas and Alvarez, 2010).
The SCSA® test is performed using a laser-based device called a flow cytometer. Essentially, the flow cytometer measures 250 sperm/s with high precision, providing a statistically robust result based on data from 2x 5000 sperm per sample. The precision of flow cytometry coupled with the precise biochemistry of the SCSA test provides highly accurate measurements of sperm chromatin and DNA integrity.
The cells are stained with a fluorescent DNA stain (acridine orange) and then forced through a glass channel in a liquid suspension. When the cells pass the laser beam, the laser light causes the dye to emit fluorescent light of a specific color.
In the SCSA® test, due to the unique nature of the Acridine Orange dye, sperm that emit green fluorescence have undetectable levels of fragmented DNA, while sperm that emit yellow to red florescence have moderate to high levels of fragmented DNA.
The SCSA test is a two-fold simultaneous flow cytometry measure of
in thousands of sperm in a fresh or frozen/thawed semen sample.
The extent of DNA strand breaks and abnormality of chromatin structure are related to male factor fertility, including time to couple pregnancy, IVF embryo quality, miscarriage, or infertility.
Male fertility is classically addressed by semen tests that include sperm density, motility and morphology. However, numerous studies over the past half century clearly show that, except for absence of sperm, these parameters do not predict pregnancy since fertile and infertile men have overlapping values. The classical semen tests are light microscope measures of external sperm factors while the SCSA test are measures of the internal nuclear factors not visible by light
microscopy.
The SCSA test is a high precision test that requires a flow cytometer which due to the need of a high capital expense and a trained technician is not amenable to most infertility clinics. Thus, semen samples collected, as per detailed instructions, in a clinic or in a patients’ home, can be flash frozen in a LN 2 dry shipper tank.(See support protocol 1).
Clinics can then send us an email informing us. A pre-cooled LN 2 shipper is then sent to the clinic and the semen samples are flash frozen and the LN 2 shipper then returned to a SCSA diagnostic testing lab.
Some clinics send frozen semen samples in well insulated dry ice containers.
SCSA clinical data are then sent via a secure WEB site to the physician or patient that ordered the test.
There are five main populations of sperm identified by the SCSA® test:
the percentage of sperm with immature chromatin. HDS sperm exhibit less chromatin condensation, resulting in increased DNA stainability. Some studies have shown that if the %HDS is above 36%, no pregnancies have occurred (Menezo)
Our SCSA clinical report lists four statistical categories of fertility potential. These were derived from a comprehensive study of male fertility potential without medical intervention and have been confirmed in additional studies.
< 15% DFI = Excellent to good pregnancy outcomes (excluding female infertility factors)
> 15% to < 25% DFI = good to fair pregnancy outcomes
> 25% to < 40% DFI = fair to poor pregnancy outcomes
> 40% DFI = Very poor pregnancy outcomes
Furthermore, the finding shows a graphical representation of the %DFI (DNA Fragmentation Index) and the %HDS (high DNA stainability)
The graph shows that 76% of the measured sperm shows no DNA fragmentation and 24% shows medium-high DNA fragmentation (DFI= 24%). Pregnancy naturally or with the help of an IUI is possible.
Current data shows that the probability of a successful pregnancy is significantly lower when the proportion of sperm with fragmented DNA is > 25%. Nevertheless, a DFI > 25% does not rule out a normal pregnancy. The 25% threshold is a statistical threshold. Therefore, if a man has a constant DFI >25%, he will be placed in a statistical group of men who have been shown in clinical trials to take longer to achieve natural pregnancy and more IVF cycles. An increased rate of spontaneous abortions is associated with this or pregnancy does not occur at all.
The ovum is believed to be able to repair a certain amount of DNA damage.
Moderate to high levels of DNA fragmentation (DFI) found in the red fluorescent sperm likely exceed the egg’s DNA repair capacity.
Marcos Meseguer et al found that when oocytes from patients were taken, for every 10% increase in DFI, there was a 1.31 times increase in probability of not achieving pregnancy. Whereas, the same was not seen when donor oocytes were employed suggesting the higher capability of young oocytes to repair sperm DNA damage.
REPEATED IUI / IVF / ICSI FAILURE : The reason behind the unsuccessful attempts at IUI/IVF/ICSI can be identified and managed
REPEATED MISCARRAGE : An explanation for repeated spontaneous abortions (RPL, repeated miscarriage) could be found and guided
POOR EMBRYO DEVELOPMENT AND QUALITY : The reason behind poor embryo quality could be the fragmented sperm DNA which could be corrected before the next treatment
MEN AT RISK FOR HIGHER OXIDATIVE STRESS:INITIAL SCREENING TEST : Especially for couples who have been attempting pregnancy for a long time to identify the best management strategy
UNEXPLAINED INFERTILITY : Couples desiring children with all normal diagnostic tests
In cases of high DFI, we suggest:
When a risk factor is identified, after avoidance / correction of the risk factor a repeat SCSA test after 3 months is advised.
Yes. We offer a SCSA test at half the cost for controlling the effect of avoidance / correction of a risk factor.
Please find a few examples below:
After quitting smoking for 84 days
3 months after varicocele correction surgery :
3 months after stopping pesticide exposure:
After 24-48 hours of abstinence, the samples should be taken by masturbation into sterile plastic sample containers. After liquefaction of the sample, the sperm concentration, motility and morphology are determined.
Cryopreservation should take place no more than 1 hour after delivery, since the DFI can be falsely raised after a longer waiting period.
Flash-freezing: Freeze 0.25mL or more of liquefied native ejaculate in 1-2mL cryovials. Cryoprotectants is not required! (Please freeze 2 cryovials per patient)
Samples should be frozen as soon as possible after liquefaction to minimize exposure to air, which can cause damage caused by reactive oxygen species.Cryovials should be tightly capped, labeled with two identifiers (e.g. name and date of birth) and placed in an upright position directly into the pre-chilled cryo-container. The container must be closed quickly.
Note: Cryoprotectants are not needed since quick-frozen sperm give the clearest SCSA® data. This feature is unique to mammalian sperm cells due to the highly condensed, crystalline nature of the nuclei.
Note: If the sperm concentration is adequate, less than 0.5 mL (two 0.25 mL aliquots) of semen may be sufficient for the SCSA® test. To perform the test, we require two 0.20 mL aliquots with a concentration of >0.5 million/ml. Please contact our laboratory if you have questions pertaining to the volume necessary for a sample of a certain concentration.
We ship a pre-chilled dry shipper. Dry shippers store the liquid nitrogen in porous material and thus ensure the safe transport of your cryopreserved samples in the nitrogen gas phase.
The low temperature in the container can be maintained for 7 days. It is therefore important to send it back as soon as possible using the mailed/supplied shipping label.
Note: liquid nitrogen is adsorbed by special materials inside the shipping container. Even if they are turned upside down during transport, no liquid nitrogen will leak out. Dry shippers are not considered dangerous.
After receiving the sample(s) in our laboratory, we need 4 working days to create a report, which we will send to you.
The cost of the test is 295 Euro.
* Samples are shipped in a pre-cooled shipper that we can provide either at ambient or pre-chilled temperatures. The pre-cooled shipper will be sent to the centre where you have provided your sample for the centre to ship it to us.
The test at present is not covered by the insurance.
Please write us an email to contact@scsalab.com and after contacting you, we will send toy a dry shipper for the patient sample to be sent to us.