Sperm Chromatin Assay

I asked my good colleague Dr. Evenson who developed the SCSA sperm chromatin/DNA test to write an info page on this very important test. Our US government uses this test to track studies in OSHA (work safety); DOD (Gulf War Syndrome studies); EPA (environmental safety); also March of Dimes has funded Dr. Evenson's work for years.

I will tell you straight up that some RE's and ObGyns don't like the test b/c some couples with high sperm DNA damage choose to not do $$$ IVF or ICSI (so they loose business). Take their recommendations with the knowledge that it isn't based on good science. Many many clinics (most good andrology ones) do use this test. You can order it with any doctors signature, even your GP!).

What exactly is the Sperm Chromatin Structure Assay (SCSA)?
The SCSA is an assessment of sperm DNA fragmentation that identifies men with highly reduced probability of initiating and supporting a successful pregnancy. The percentage of sperm in a semen sample with fragmented DNA is reported as the DNA fragmentation index (%DFI).

What will the SCSA tell me?
The SCSA can help answer difficult questions regarding fertility options based on the integrity of the sperm DNA. After the SCSA analysis is completed, a Clinical Report is generated containing two numbers: 1) the %DFI (DNA fragmentation- %sperm containing damaged DNA) and, 2) %HDS (% High DNA Stainability sperm with immature chromatin). The %DFI number will place the patient in one of the following three groups:

< 15% DFI = excellent sperm DNA integrity
> 15 to < 30% DFI = good sperm DNA integrity
> 30% DFI = fair to poor sperm DNA integrity

It is important to note that a DFI value above 30% does not preclude a normal, full term pregnancy. A >30% DFI, if consistent over time, does mean that the male partner is statistically placed into a group of men that demonstrate a longer time period to establish a pregnancy, more IVF/ICSI cycles, increased risk of spontaneous abortions or no pregnancy.

My semen analysis showed normal levels of sperm concentration, motility and morphology. Can I still have high levels of DNA fragmentation?Yes. Sperm that appear to be completely normal by all the standard measurements may have high levels of DNA fragmentation.

What are some conditions that might indicate the need for an SCSA test?*abnormal semen analysis
*unexplained infertility
*persistent infertility after treatment of female
*recurrent miscarriage
*prior to assisted reproductive technologies
*cancer in male: before and after treatment
*advancing male age, e.g., ~ age >45
*varicocele

How can the SCSA help me and my physician?
A recently published meta-analysis for SCSA data and pregnancy outcomes showed that couples have a 6.5-10 times, 7.1 – 8.7 times, ~2 times, and ~1.5 times greater probability of a successful pregnancy by natural, IUI, routine IVF and ICSI fertilizations, respectively, if the semen samples contains < 27-30% DFI.

The in-vivo and IUI meta-analyses were similar, indicating that IUI infertility patients with <30% DFI have as good a statistical probability of obtaining a pregnancy/delivery as presumably fertile couples trying to become pregnant with the same %DFI.

What causes DNA fragmentation?  DNA fragmentation in sperm may be the result of many factors including, but not limited to, disease, diet, drug use, high fever, elevated testicular temperature, air pollution, cigarette smoking, some prescription medications, varicocele and advanced age. In about 50% of infertile couples, male-factors play a significant role in infertility.

What are physicians saying about the SCSA?  Current conclusions of an international review by Erenpreiss et al. (2006) state: “Without doubt the existing data justify the necessity to introduce sperm DNA damage assessment into the routine infertility investigation. The SCSA is currently the only method that has provided clear clinical cut-off levels and that can be recommended for a robust sperm DNA damage evaluation%”.

My physician requested that I have the SCSA test and gave me the telephone number for SCSA Diagnostics, Inc. What do I need to do?  SCSA Diagnostics can send the sperm shipping container and all of the necessary supplies to your physician's office or directly to your home. The sample collection, packaging and shipping procedures are simple and completely explained on the SCSA website (www.scsadiagnostics.com) as well as in the instructions provided. Patients typically have very few questions and do not make mistakes. The results of your SCSA analysis are sent to your physician within one week of receiving your sample at SCSA Diagnostics.

Dr. E

Sperm Leaking Out After Intercourse- Lessons in Sperm Transport Through the Cervix

Q: Why does sperm leak out sometimes and not others? Does it mean the cervix is closed? Or there is more sperm sometimes than other times? If a lot leaks out, does that mean there was a lot of sperm? Is there any way to prevent it from leaking?

A: In a 5 yr study of 11 women (Baker & Bellis, 93), sperm loss after intercourse ("flow back") was observed- Flow back occurred 94% of the time, with an average loss of 35% of the sperm. It is totally normal and is not a sign that there is anything wrong. It is more pronounced the larger the ejaculate volume, and remember ejaculate quantity is impacted by "how turned on" your husband is. So if you have a great session, or it has been awhile since doing the deed, there will be more.

The sperm that penetrate into the cervical mucus begin to do so within 1.5 min, and they are pretty much done by 30 minutes, with no gain in sperm numbers in the cervical mucus or Fallopian tubes after 45 min from intercourse. Only thousands of the millions of sperm ejaculated in the vagina make it to the cervix and only hundreds of these make it to the Fallopian tube!

The very best of the best get there, the rest get washed out- it is OK!

Dr. E

Sperm Morphology. Please help me understand

Q: I have talked to my RE again regarding my DH's SA results. He said that he did not feel it is necessary to do a second SA for my DH. He scheduled me for a consultation on March 15th. It is really a long time and will be after I again. Another wasted cycle.

Since I have done all the blood tests (P4) etc.. I asked his office for a print out of my results. I have checked all my numbers - they are all normal. I have also asked for a print out of my DH SA. I still think that his low morphology is an issue.

So here is a complete SA:
Liquefaction: Normal
Color: Normal
Volume: 2.6
Viscosity: Normal
Agglutination: None
Sperm Count : 117
% Motility: 81%
Progression: Good

Tygerberg Strict Morphology
% Normal: 1%
Comments: few w/coiled tails, accelongated heads

I am looking for your opinion. (My HSG, Blood work - all is good). In addition, what questions should I ask my Dr. when I see him in March?

Thank you for your time!
7 years 6 months 17 days 2 hours 51 minutes since our wedding day

A: Please see my FAQ on RE's versus Andrologists. You know you are working with a good andrology laboratory IF they follow the World Health Organization guidelines Standard Procedures for semen analysis, which state "Two samples should be collected for initial evaluation". Almost 100% of the time the first sample your DH provides for an SA will be of a poorer quality than subsequent samples. In fact, so much so, that for my NIH studies using normal fertility donors, I CAN'T EVEN USE THEIR FIRST EJACULATES, b/c they are such poor quality and these are normal fertile guys!

I hate to say this, but...if a clinic only goes on one ejaculate then they will have a higher percentage of couples that need extreme interventions such as ICSI...if you get my drift.

That said your DH sounds like he has some real issues, but I would INSIST on a second SA or get one done elsewhere so you know for sure what you are dealing with. His count and motility are good, so I would love to have a true andrologist look at his sperm and help you know what kind of defects he has etc... And I would use a collection condom, you are probably trying to find out if you are providing a sample that can be used for IUI, IVF or ICSI and the condom will maximize the normal sperm he can produce-- so you can better decide which procedure to use.

I have asked a colleague of mine who was part of the panel to WRITE the WHO guidelines to summarize morphology for all of us (below)!

Don't be afraid to be pushy with your docs! It is your money and your future baby You should also ask what percent of their IVFs are done by ICSI again see the RE FAQ-

Finally, get DH on a fertility vitamin for sure, as these do improve morphology in some men!

SPERM MORPHOLOGY
The "normal range" for human sperm morphology, i.e. the proportion of sperm that should have normal morphology, has evolved enormously over the past 50 years. It used to be that semen analysis was judged on "the rule of 60s", i.e. 60 million per milliliter, with 60% motile and 60% normal, but that changed in a very informal way to a more generally accepted 20 million per milliliter and a lower expectation of normal forms. But even though sperm count (we don't use the word "density" as that can get people confused between how dense sperm are (i.e. mass per unit volume) and what we're really talking about, which is their concentration), can be measured reasonably accurately even by non-expert labs if they follow the right rules (although a lot just don't get it right), sperm motility is rather more difficult as the human eye is attracted to moving objects, and so, when looking down a microscope, all but the best-trained technicians have a tendency to over-estimate. But when we come to sperm morphology the situation is astonishingly variable.

Deciding which sperm are "normal" is highly subjective and, despite great efforts by organizations such as the WHO, cannot be standardized by reading from a book. "Hands-on" training (perhaps it should be "eyes down" training?) is essential. In 1992 the WHO attempted to make sperm morphology a lot more standardized by telling people in the 3rd edition of its Lab Manual that they should be more strict, and that "borderline" sperm must be considered as abnormal , since it's always easier to standardize something that's strict compared to something that's more wishy-washy. However, about the same time, a highly standardized scheme of "strict morphology" was being promoted around the world, coming from the Tygerberg Hospital in Cape Town, South Africa. The popularity of this "Strict Criteria Morphology" system grew out of the close collaboration by a young clinical researcher at Tygerberg, Dr Thinus Kruger, and the Jones Institute at Norfolk, Virgina, hence the common name for this scheme is "Kruger morphology". But the scheme was actually developed by Dr Roelof Menkveld, the scientist running the sperm lab at the Tygerberg Hospital, and hence the scheme should be referred to as the "Tygerberg Strict Criteria" (or
TSC").

Anyway, there was confusion between the 1992 WHO "stricter" scheme that had a supposed normal limit of 30% normal forms and the TSC which had cut-offs of 14% for poor prognosis (at IVF, not for in-vivo fertility at that time) and 5% below which ICSI was recommended. A careful collaborative study between Dr Menkveld's lab and that of Dr David Mortimer in Sydney, Australia (who had written that part of the 1992 WHO Lab Manual) revealed that the two schemes, rather than being in competition, were actually very similar, if the WHO scheme was applied as had been originally intended! The 30% cut-off in the WHO Lab Manual was actually a "guesstimate" for the new scheme decided by a committee, and was to have been backed up by research studies – but they never happened due to funding cuts at the WHO.

There is also the important aspect that some people have false impressions of what sperm morphology means. It's not a "beauty pageant": sperm shape is extremely variable between different species, so if a long thin head was an absolute abnormality then some species would never be able to breed. And anyone who has seen the huge, hook-shaped heads of rodent sperm knows that if such shapes were seen in a human sperm morphology slide, the man would be considered to have terribly abnormal sperm – but they work just fine for rats and mice. And yet, even what's normal for a mouse would be abnormal for a rat, and vice versa!

And of course, there is no absolute relationship between normal/abnormal morphology and a sperm's ability to fertilize or not. Furthermore, there's no actual relationship between abnormal sperm morphology and abnormal embryos (or babies!): scientists call this then "phenotype" of the sperm not being directly related to its "genotype". Morphologically abnormal sperm can sometimes fertilize – and "normal" sperm probably quite often have other defects that just couldn't be seen under the light microscope (or were biochemical abnormalities, not structural ones) that prevent them from fertilizing.

So what does a sperm morphology result mean? Basically, any assessment of sperm morphology – even one performed by a highly skilled, fully-trained technician or scientist working in a lab with sound quality control systems – is no more than an assessment of how well the process of sperm production seems to have worked. There are many opportunities for there to be finer defects in sperm than can be seen in stained preparations under the light microscope, and so when a sperm is described as being "normal" there is real presumption that it is, in fact, normal at all! All sperm morphology assessments are just simply indicators of how serious the risk might be that the sperm won't work, whether it be when insemination occurs during sex, or by IUI, or by IVF (sperm morphology is generally accepted to be irrelevant for fertilization using ICSI). There are also issues of accuracy and reliability of sperm morphology assessments when only 100 (or 200) sperm are examined: statistical sampling error must be acknowledged, and this says that when based on on such counts, values of even 3% and 8% are not necessarily actually different.

So with a value of less than 5% normal forms there is only a higher risk that IVF won't work, and that ICSI should be used. With 5% to 14% normal forms IVF is likely to be needed to improve the chances of conceiving, although IUI can be quite successful with less-severe cases. So making a distinction based on a measurement with a very limited "dynamic range" can be difficult, and that's why some specialized sperm labs used other information as well. Dr Menkveld has another measurement called the "Acrosome Index" that helps identify men who will have a serious problem achieving fertilization even by IVF, and the WHO has the "Teratozoospermia Index" (or "TZI"), which is a measure of how abnormal the abnormal sperm are. A high TZI (above 1.8) indicates that there is a great likelihood that even those sperm that were seen as "normal" under the light microscope will be unable to function properly.

In conclusion, like every aspect of a semen analysis, a sperm morphology evaluation is not a "black-and-white" assessment, a poor result only indicates an increased risk of more severe male factor subfertility.

Dr. E

Sperm Transport to the Fallopian Tubes

Q: If a man has normal sperm count and normal sperm motility, approximately how many sperm will be able to get to the fallopian tubes for each ejaculate during a woman's ovulation period? I've read different information on this. Some said about 1000 sperm to 5000 sperm but some said only about 50 sperm to 200 sperm.

A: This is actually a very good question, one only a handful of scientist around the world (including myself) have spent our careers studying.

Here is the issue-- Prior to the 1990's several good studies were done looking at sperm in the tubes after insemination, by flushing the tubes with salt water and counting the sperm. But back then we hadn't done the studies I did on my PhD, (along with others) to show that sperm actually bind or stick to Fallopian tube cells, and that they stay stuck even if you rinse the tube. These stuck sperm are released in waves over time, so there will always be sperm available to meet the egg. Once sperm are released from the tubal cells they either meet an egg or they die within a few hours as they are "capacitated". This gives a supply of ready sperm as they wait for the egg for days to a week plus.

These early studies showed really low sperm numbers in the tubes- 1 out of every 14 million inseminated sperm got inside the tube, and only 1 of every 2000 sperm inseminated made it into the cervical mucus (Settlage et al 1973).

Likely, there were other sperm in the tubes in these studies that were attached to the tubal cells and therefore, not counted. Since the 1990's though the kind of studies you can do in people have drastically changed. It is almost impossible to get permission to have a couple have unprotected intercourse at ovulation, and then surgically remove a portion of the tube to study for counting sperm attached to the tubal cells. You could be setting up a tubal pregnancy.

For this reason, when I had my tubal ligation I took estrogen (to mimic ovulation), had intercourse and then 24 hrs later had a tubal and my tubes removed to look by scanning micrograph for the sperm attached to the tubal cells. I found about 20 sperm in the portions of the tube I looked at. Just call me Madame Currie!

So... no one knows the answer to your question for sure. In animals, where we have done many studies -- only hundreds of sperm are in the tube (even though their ejaculates can have up to 350 million sperm in them).

One other thing I have found is that with stallions, the number of sperm that attach to tubal cells in culture (laboratory) is very, very highly correlated to their ability to impregnate mares (r=0.80). The stallion is a great model b/c they may breed 20-60 mares a season so you can get good pregnancy data on more than one female, and compare that to sperm function.

In men, I have seen a wide range of ability of sperm to attach to tubal cells in culture as well. Some men have almost no sperm attaching and they die quickly within hours, while some men have a very high attachment rate and survival rate with sperm living attached to the tubal cells for up to 9 days in the laboratory. This correlates with what has been in seen in women with live sperm found in one woman’s tubes 21 days after intercourse!

Extrapolating from the well done stallion study, we can assume that the men with the most sperm attaching to the tubal cells and therefore living the longest, will have the highest fertility.

Thanks for asking a fun question!

Dr. E