Investigating unexpectedly positive hCG test results


In February, I wrote about interfering antibodies being the cause of false-positive hCG results from blood samples.  As a follow-up, I thought it would be a good idea to talk about what the lab can do to investigate the potential problem.

As a reminder: false-positive hCG results are often attributed to the presence of an interfering antibody in the blood sample.  Such was the case with Jennifer Rufer who was misdiagnosed with cancer due to a false-positive hCG blood test.  It's important to know that laboratories are likely not able to independently identify when an interfering antibody is present.  An interfering antibody should be suspected whenever the clinical picture of the patient fails to match the laboratory (in this case, hCG) results.  When laboratorians are asked to investigate the possibility of a false-positive hCG result there are several ways to do so but the key piece of information to note is that the lab has to be notified that there is the suspicion of an erroneous hCG result.  This is because the laboratory is nearly always unaware of the clinical picture of the patient.  We need to rely on our clinical colleagues to alert us to the possibility that an hCG test result is potentially incorrect.

So, what can the lab do when asked if an hCG blood test result is falsely increased?  There are several approaches the lab can take to investigate:

  1. Perform an hCG test on a urine sample obtained from the patient.  Because hCG is excreted in the urine, the detection of hCG in a urine sample indicates that the result from the blood sample is likely accurate.  Interfering antibodies aren't excreted into the urine and so if hCG is detected in the urine then the blood test result is probably really due to the presence of hCG.
  2. Perform a dilution of the blood sample.  Because interfering antibodies are reactive against the hCG assay reagents, the expected response to sample dilution is typically not observed.  That is, if the sample is diluted by a factor of 1:2, the hCG concentration should decrease by a factor of 2 if the hCG molecule is truly present.  Failure to observe the expected decrease in concentration supports the presence of an interfering antibody.
  3. Repeat the hCG test using a different method.  Interfering antibodies may be reactive against antibodies derived from a specific animal species (e.g. against mouse antibodies).  If the hCG test is repeated using a different method and the results are considerably different then that can suggest the presence of an interfering antibody.  Importantly, the alternative method selected should be one that uses antibodies derived from a different species of animal than the test in question.
  4. Treat the sample with blocking agents.  Blocking agents are commercially available that can be used to remove (by adsorption) potentially interfering antibodies from the sample.  If the hCG results after treatment is considerably different from the result of the untreated sample, then interfering antibodies may be present.  The definition of considerably different is not well defined although many labs use a difference of 50% to alert them to the possibility that interfering antibodies are present.

I recommend that laboratories use more than one of these investigations rather than rely on a single one.  As the Rufer case so clearly demonstrates, erroneous laboratory results can result in serious harm to patients.  Together with our clinical colleagues, it our responsibility to do all that we can to assure that laboratory test results can be correctly interpreted.

    Choosing a screening test for Down syndrome


    Last month I wrote about the many different tests used for Down syndrome screening.  Today, there are 6 different screening tests that a woman can choose from should she opt for any testing at all.  Selecting one can't be an easy task for most women and, given the relatively short time that most physicians have to spend with patients today, many simply follow their doctor's advice and don't truly understand their options.

    The choice of which Down syndrome screening test to have depends on the answers to a few questions:

    • When during pregnancy does the patient first seek obstetrical care for her pregnancy?

    Some screening tests are performed in the first or the second trimester while other tests are performed in both first and second trimesters.  If a woman first seeks obstetrical care after the 14th week of pregnancy then only the second trimester tests can be offered.  That means either the Triple or the Quad tests.  If she presents earlier then all screening tests can be considered.

    Nuchal translucency (NT) is an ultrasound measurement of space under the skin behind the fetal neck and is performed only during the first trimester.  An increased NT is strongly associated with Down syndrome and other chromosomal abnormalities.  It's a component of the Combined test, the Integrated test, and the Sequential test.

    Chorionic villus sampling (CVS) is the removal of a small piece of placental tissue in order to obtain fetal cells in order to obtain the fetal karyotype.  The karyotype will identify the number of chromosomes the fetus has inherited and is usually only performed after an abnormal Down syndrome screening test.  This procedure is performed in the first trimester because amniocentesis can't be performed this early in pregnancy.

    If CVS is not available then options include the Integrated, the Triple, or the Quad test.  If the NT is not available then the choice of screening tests is limited to the Serum Integrated test, the Triple test, or the Quad test (the same is true if both NT and CVS are not available).

    • When does the mother want the results of the screening test?

    If the woman wants the earliest assessment of the risk of carrying a fetus with Down syndrome then she should opt for the Combined test.  The test is performed only in the first trimester and results are available sooner than with any of the other test options.  However, the test doesn't perform as well as some of the other tests.  On average, the Combined test detects about 85% of all Down syndrome fetuses at a 5% false-positive rate.

    If she's willing to wait until the second trimester for the test results then the best screening test is the Integrated test.  This test gives the highest Down syndrome detection rate (about 95%).  The test can also be performed without an NT (the Serum Integrated test) and will provide the same detection rate with a slightly higher false-positive rate.

    If the patient desires early risk assessment but is willing to wait for test results if needed, then the Sequential test may be appropriate.  The Sequential test reports results in the first trimester only if the risk of having a Down syndrome fetus is very high.  If the risk is not high, then results are not reported until the second trimester test is completed.

    The table below summarizes the performance of the 6 different Down syndrome screening tests.  The "detection rate" (DR) is the percentage of Down syndrome pregnancies that are correctly identified by the test and the "false positive rate" (FP) is the percentage of unaffected pregnancies identified as abnormal.

    Performance of maternal serum screening tests

    Selecting a test for Down syndrome screening doesn't have to be complicated.  Patient's should be informed about the pros and cons of each test (and screening tests in general).  It's often said that information is power and, when it comes to Down syndrome screening, women should be empowered to choose the test that is right for them.

        Noninvasive antenatal testing using cell free fetal DNA


        Today's post is by a guest author, Joshua Cook, M.D.  Dr. Cook is a pathology resident at the University of Utah.

        Fetal cells, largely of placental origin, circulate freely in the mother’s bloodstream during gestation.  In some cases fetal cells are detectible in maternal blood for years post delivery.  Cell free fetal DNA (cffDNA) is also present in the peripheral blood of pregnant women, though it disappears rapidly at delivery.  cffDNA appears early in the first trimester and tends to increase in concentration as gestation progresses.  A number of studies over the last decade have shown this naked fetal DNA provides a noninvasive method for rendering prenatal diagnoses without risk to the fetus or mother.

        Consider hemolytic disease of the newborn (HDN).  About 10% of pregnancies in Caucasian RBCs populations involve a rhesus Rh(D)- negative mother carrying an Rh(D)-positive fetus.  Such a mother is at risk of becoming sensitized, or isoimmunized, to Rh(D)-positive red blood cells following exposure to fetal blood. 

        Small physiologic fetal-maternal hemorrhages commonly occur at delivery or during miscarriage.  Exposure of the mother’s immune system to fetal red blood cells may result in the generation of maternal antibodies to fetal antigens (sensitization or isoimmunization).  These antibodies may then cross the placenta in subsequent pregnancies and harm a future baby.  In pregnancies with Rh(D) incompatibility [Rh(D)-negative mom with Rh(D)-positive fetus] the mother has historically had about a 10% risk of becoming sensitized. 

        Knowledge of the fetal RHD genotype in Rh(D) negative women greatly simplifies antenatal management.  Modern medical practice requires that immunoprophylaxis (anti-D IgG immunoglobulin; aka RhoGAM) routinely be given to all non-sensitized RhD-negative woman in pregnancy to prevent isoimmunization.  This had led to a dramatic decrease in HDN since the 1960’s.  When administered, the anti-Rh(D) antibodies circulate in the mother’s bloodstream and prevent her immune system from reacting to Rh(D)-positive fetal blood.  The standard of care is to provide anti-Rh(D) administration at 26 to 28 weeks of gestation, and after suspected antepartum fetal-maternal hemorrhage (i.e. amniocentesis, chorionic villous sampling, pregnancy loss, ectopic pregnancy).  Anti-D IgG is also given with within 72 hours of delivery if the baby is found to be Rh(D)-positive.

        Because the Rh(D) status of the fetus is usually not known prior to birth, about 40% of immunoprophylaxis is given unnecessarily to Rh(D)-negative mothers who are carrying Rh(D)-negative fetuses.  cffDNA testing restricts immuneprophylaxis to the 60% of pregnancies with a true Rh(D)-positive fetus.  Fetal RHD genotyping using maternal plasma is about 95% accurate at 11 -13 weeks gestation and is nearly 99.8% accurate by 26 weeks (when the first dose of immuneprophlyaxis is administered).  Accuracy improves throughout gestation because the amount of cffDNA in maternal peripheral blood increases with gestational age. 

        cffDNA testing is currently used in Europe to identify high risk pregnancies [i.e. Rh(D)-positive fetus  in a previously sensitized mother] which benefit from intensive antenatal monitoring.  cffDNA RHD genotyping is used to determine the risk of chorionic villous sampling (CVS) in sensitized women.  CVS is a technique used in the prenatal diagnosis of fetal aneuploidies (such as Down syndrome) and other inherited genetic disorders; however it is relatively contraindicated in isoimmunized women since procedure related fetal-maternal hemorrhage exacerbates pre-existing sensitization.  If the fetus lacks Rh(D) then boosting the maternal amnestic response is not an issue.

        Cost and outcome studies for noninvasive cffDNA prenatal screening are in progress, but obstetricians may soon have a powerful new tool in their armamentarium.

         

         

        Fetal lung maturity tests. Are they truly necessary?


        When infants are born before 39 completed weeks of gestation, they are at increased risk of developing respiratory distress syndrome (RDS).  The risk of RDS increases as the gestational age of the infant at delivery decreases.  In other words, the more premature a baby is born, the more likely it is that it will have RDS.

        This is a primary reason why an elective delivery before 39 weeks of gestation should not take place unless the fetal lungs are shown to be mature using fetal lung maturity tests.  The logic behind that mandate should be obvious: infants that develop RDS can die and those that do survive often develop other serious complications like septicemia,  necrotizing enterocolitis, retinopathy, and developmental handicaps.

        Intuitively, it makes sense that if lab testing demonstrates mature fetal lungs then the risk of RDS, and its associated complications, would be low and elective delivery before 39 weeks could be permitted.  However, a recent study showed that even after documented fetal lung maturity, infants born before 39 weeks were at higher risk of adverse outcomes than infants born at 39 to 40 weeks.

        Infants born before 39 weeks were, overall, at 1.6-fold greater risk of having something bad happen to them.  Things like elevated serum bilirubin, ventilator support, low blood glucose, admission to a neonatal intensive care unit, or even RDS!

        Results like these definitely call into question the current convention that delivery before 39 weeks is okay if fetal lung maturity is confirmed by lab testing.  One has to even wonder why fetal lung maturity tests are even necessary.  Perhaps they aren't.

        Due to improvements in gestational age dating, maternal administration of corticosteroids that accelerate fetal lung maturity in at-risk pregnancies, and exogenous surfactant replacement therapies, the number of newborn deaths due to RDS has declined considerably over the last 15 years.  Also, doctors are ordering fewer fetal lung maturity tests than they have in the past.  They probably aren't going to go away any time soon but all signs are pointing towards their demise.

        A growing body of evidence is telling us that 1) elective delivery of infants before 39 weeks of gestation should be avoided; 2) that's true even if their lungs are shown to be mature by lab testing; and 3) testing for fetal lung maturity is decreasing.

        Perhaps it is time to send these tests away once and for all.

        A perfect Down syndrome screening test?


        In a January post, I wrote about a possible new Down syndrome screening test that detected 100% of affected fetuses and had a very low false-positive rate.  Recently a different group of scientists in Cyprus reported they had developed a perfect test to detect Down syndrome: one that correctly detects all Down sydrome fetuses but had zero false positive results.

        While the earlier report utilized a time-consuming and expensive technique called massively parallel sequencing, this new study focused on a process called DNA methylation.  Methyl groups are chemical structures that are naturally attached to regions of DNA.  They function to turn genes on or off.  Because DNA from fetuses have different methylation patterns compared to their mothers', fetal DNA can be distinguished from mom DNA in a blood sample taken from the mother.

        After enrichment of the methylated fetal DNA from the mother's blood, quantitative PCR was used to amplify specific methylated regions of chromosome 21.  Fetuses with Down syndrome were identified because they have three, rather than two, copies of chromosome 21, and the increased copies were readily detected by the test.  Out of 40 women who were 11 to 14 weeks pregnant when tested, the method was 100% accurate: all of the 14 Down syndrome and all of the 26 unaffected fetuses were correctly identified.

        In contrast to a sequencing approach, the techniques used in this study are rather easy and fast to perform and don't require expensive equipment or software.  It is also a lot cheaper.

        This is a promising test but there's still a lot more to be done before it makes the leap into clinical use.  The Cyprus researchers are planning to do larger-scale clinical trials with many more women.  Let's hope that the results from those trials are as exciting as this early report.

        Why so many Down syndrome screening tests?


        Throughout pregnancy, women have to make lots of decisions.  Two of those are “Do I want to have a screening test to see if my baby has an increased risk of Down syndrome?” If the answer to that is yes then the logical next question would be “What tests can I choose from?”  That is not an easy question for most women to answer without some guidance from their doctors and it’s not easy for many doctors to help provide that guidance.  Why?  Simply put, there are many tests from which to choose.

        Let’s be clear on an important concept up front.  Screening tests are not diagnostic tests.  A test that screens for Down syndrome doesn’t identify if a baby has Down syndrome; it identifies babies that are at increased risk of having Down syndrome.  Women with abnormal or positive screening test results can undergo additional tests that can be used to confirm if their baby does or does not have Down syndrome but the screening tests cannot do that. 

        All Down syndrome screening tests require a blood sample from the mother.  Biochemical markers in the blood are measured in the laboratory and the results used to calculate the risk that the baby has Down syndrome.  Those biochemical markers include:

        • Alpha-fetoprotein (AFP)
        • Human chorionic gonadotropin (hCG)
        • Unconjugated estriol (uE3)
        • Dimeric inhibin A (DIA)
        • Pregnancy-associated plasma protein A (PAPP-A)

        Some screening tests also include the measurement of nuchal translucency (NT) that is obtained by an ultrasound scan of the fetus.  The NT is the width of the space between the spine and skin at the fetus’ neck.

        All together, these 5 biochemical markers and 1 ultrasound marker can be used in various combinations to create the different Down syndrome screening tests.  There are 6 to choose from:

        Table

        Some, like the Triple (3 markers) and the Quad (4 markers) tests are performed on a blood sample collected during the 2nd trimester and don’t require the NT ultrasound measurement.  The Combined test (so called because it “combines” the biochemical and ultrasound tests) is performed only in the 1st trimester.  All the other tests use two different blood samples (one collected from the mother in the 1st and the other in the 2nd trimester) and may or may not also include the NT measurement.  No wonder this is confusing!

        Why are there so many different tests?  Two reasons: first because Down syndrome screening tests evolved over many decades (and continues to evolve) and second because the medical community is often slow to change its habits.

        In the not so distant past, one “test” was used to determine Down syndrome risk: the age of the mother.  We actually still use the mother’s age in determining Down syndrome risk.  The risk of having a baby with Down syndrome increases as the age of the mother increases.  We now use this age-based risk as a starting point; a risk that is the modified by the results of the screening test.

        In 1988, the Triple test was introduced as a way to adjust the age-based risk using the measured concentrations of AFP, hCG, and uE3 in the mother’s blood.  A few years later, the Triple test turned it into the Quad test when it was discovered that the addition of DIA to the Triple test improved the Down syndrome detection rate.  However, the Triple test didn’t disappear and labs simply continued to offer it as well as the Quad test.

        The Combined test appeared in 1999 and had a Down syndrome detection rate that was similar to that of the Quad test.  Unlike the Quad test, however, the Combined test provided women a way to get Down syndrome screening test results many weeks sooner.  So, it was added to the menu along with the Triple and the Quad tests.

        It wasn’t long before someone thought to “integrate” the Combined test with the Quad test and thus was born the Integrated test.  And, for those women without access to the specialized equipment needed to perform the ultrasound NT measurement, the Serum Integrated test required only the mother’s blood samples to be tested.  While both of these tests provide the greatest Down syndrome detection rates, the need for the second blood sample collected in the 2nd trimester means that early risk-assessment is not possible.

        And finally, the Sequential test developed as a modification of the Integrated test.  While the Integrated test delivers its results in the 2nd trimester after all testing has been completed, the Sequential test offers women results in the 1st trimester only if the risk of Down syndrome is very high.  In the absence of a high risk, results are provided only after the test is completed in the 2nd trimester.

        Because it’s a test with a lot of history and experience behind it and because it’s well known to the doctors that order Down syndrome screening tests, the Quad test still leads the pack in terms of test usage.  The Triple test is slowly falling out of favor (as it should) and the Combined, Integrated, and Sequential tests are gaining more traction, albeit slowly.  As I said earlier, old habits are slow to change.

        Predicting preeclampsia


        Blood pressure Preeclampsia is a hot topic!  Lots of research is going on in an effort to identify biomarkers that can be used to predict preeclampsia in pregnant women.  Why all the fuss?  Well, simply put, preeclampsia is rather dangerous.  A quick review of the facts:

        • Preeclampsia is the development of hypertension (high blood pressure) and proteinuria (protein excreted in the urine) that occur in a woman after the 20th week (late 2nd trimester) of pregnancy.
        • Complications of preeclampsia include:
          • Its progression to eclampsia, which is the onset of seizures in the mother.  This occurs in about 1% of women with preeclampsia and is potentially life-threatening.
          • HELLP syndrome (Hemolysis, Elevated Liver enzymes, and Low Platelets). This occurs in 10-20% of women with preeclampsia and is potentially life-threatening.
          • Bleeding problems
          • Placental abruption (the separation of the placenta from the uterus before the baby is born)
          • Rupture of the liver
          • Stroke
          • Death (albeit rarely)
        • Preeclampsia can't be prevented and the only effective treatment is delivery of the placenta (and, obviously, the baby)

        So, if preeclampsia can't be prevented and the only effective treatment is delivery, then what good is predicting it?  That's a good question!  One reason to identify robust prediction tests is that they would permit more effective research into ways of prevent it from occurring in the first place.

        Most of the research in preeclampsia prediction tests is focused on chemicals that can be measured in the mother's blood, and that list of chemicals is very, very long.  The darling of all biomarkers studied to recently are the so-called "angiogenic" markers.  These are molecules that are involved in the growth and development of blood vessels.  Because preeclampsia is believed to be caused by abnormal placentation (and the placenta is full of blood vessels), angiogenic markers are logical candidates for predictive tests.

        I believe that the jury is still out when it comes to the actual use of angiogenic or other biomarkers for predicting preeclampsia.  Some diagnostic companies have jumped on the bandwagon to bring these tests to clinical laboratories but I think this is a case of "watch and wait."

          Can patient history and physician suspicion accurately exclude pregnancy?


          Yes, according to a study published in The American Journal of Emergency Medicine.  The goal of the study was to determine if a woman's self-assessment of her pregnancy status was reliable.  The hypothesis was that there was a very low chance of pregnancy in a woman who felt that her being pregnant was impossible.

          The study included 377 adult women of childbearing age who came to the emergency department for a variety of reasons.  These women were asked to identify their pregnancy status as "impossible," "possible," or "definite."  Similarly, the treating physician was also also asked to estimate their patient's likelihood for pregnancy as "high," "moderate," or "low."  All women had either a urine or a serum hCG test performed.

          So how'd they do?  Well, 65% of the women thought it was "impossible" that they were pregnant and they were right.  None of them had a positive hCG test.  The doctors did just as well.  None of the patients that they ranked as having a "low" chance of being pregnant were.  The study concluded that routine pregnancy testing may not be required in adult women of childbearing age.  The researchers don't go so far as to suggest that pregnancy testing should never be done if suspicion is low and caution that hCG tests should be done on patients with high-risk signs of ectopic pregnancy regardless of what the patient (or their history) suggests.

          There are a few problems with this study though.

          • First, there is no indication of which patients had a urine hCG test and which ones had a serum test.  These two tests aren't the same and urine tests probably aren't as good at ruling-out pregnancy as serum hCG tests are.
          • Second, the researchers acknowledge that all of the women in the study were seen at one suburban medical center and that they may have been confident of their pregnancy status due to the use of home pregnancy tests.  The women were also likely to be more knowledgeable about their bodies and safe sex practices.  Clearly these women don't represent the diversity of patients seen in different emergency departments.

          Still, I thought the study was interesting.  And, in an time when lab tests often seem to be used with impunity, it was refreshing to consider the diagnostic value of what the patient reports and what their physician thinks.

          False positive blood pregnancy test results


          Today I gave a lecture on hCG testing to 4th year medical students at the University of Utah.  Part of that lecture included the tragic ordeal of Jennifer Rufer.  In 2001, she was awarded nearly 16 million dollars because of a misdiagnosis of cancer from a false-positive pregnancy test.  I previously wrote about false-positive pregnancy tests from urine specimens.  This post deals with false-positive results from blood tests.

          The same things that can cause false-positive results from urine specimens can also cause false-positive results from blood samples.  It might help to review those first.  There is another cause of false-positive hCG test results that is unique to blood samples: interfering antibodies.

          First, a quick summary:

          • Human blood normally contains many different types of antibodies: molecules made by our immune system that identify and neutralize invaders such as bacteria and viruses.
          • Animals make antibodies, too, and their antibodies are used to make the components of hCG tests.  For example, mouse antibodies that recognize human hCG molecules are used in tests that measure the concentration of hCG.  One mouse antibody binds to one part of the hCG molecule (capture antibody) and a second mouse antibody binds to a different part of the hCG molecule.  That second mouse antibody has a signaling molecule attached to it.  We can measure the strength of that signal and that signal strength will be equal to the amount of hCG that it is bound to:

            True-positive hCG

            • Some people's immune systems make antibodies that react against animal antibodies.  These "anti-animal" antibodies can cause false-positive hCG test results because they connect the capture antibody and the signaling antibody together even when hCG is not present, like this:

            False-positive hCG
            This was the cause of Jennifer Rufer's false-positive pregnancy test.  Unfortunately for her, the false-positive result was recognized for what it was only after she had been treated for what her doctors thought was an aggressive type of cancer called choriocarcinoma.

            It's uncertain how many people have interfering antibodies in their blood.  Some estimate the frequency as high as 10-20%, other studies put that number much lower at <1%.  It doesn't really matter how many people have them but what does matter is how doctors and laboratorians work together to identify this problem when it occurs.  That's easier said than done.

            Why?  Because the laboratory that does the testing is nearly always unaware of the medical history of the patient so it's practically impossible for the lab to know if an hCG result is inconsistent with the patient's condition.  The lab has to rely on the doctor to question an hCG test result when it don't fit what they are seeing in their patient.  If the hCG result doesn't fit the clinical picture and the doctor calls the lab for help, there are investigations the lab can do to help identify if an interfering antibody is present or not.  But without that call questioning the result, the lab doesn't have reliable mechanisms for identifying all cases of interfering antibodies.

             

             

              The many faces of hCG


              One of the things that I find interesting about hCG is that there is more to it than meets the eye.  We often talk about hCG as though it's a single molecule when, in fact, there are lots of different variants of hCG.  Here's a quick summary of what those different molecules are all about.

              1. Intact hCG: This is the variant that gets the job done.  That is, it's the biologically active form of the hormone that is made by the placenta.  It's job is to keep the blood progesterone concentrations high which is critical for maintaining pregnancy.  It's made up of two different protein subunits simply called alpha and beta.  The alpha subunit isn't unique to hCG because it's also part of three other completely different hormones: thyroid stimulating hormone, follicle stimulating hormone, and luteinizing hormone.  The beta subunit is unique to just hCG (the other three hormones also have their own unique beta subunit).  A complete, intact hCG molecule is made when the alpha and beta subunits are attached to each other.  During pregnancy, most of the hCG in the blood is intact hCG.
              2. Nicked hCG (hCGn): Nicked hCG is made when a chemical bond in part of the beta subunit is broken.  When that happens, the hormone loses is biological activity and the alpha and beta subunits will often come apart.  During pregnancy, nicked hCG makes up about 10% of the total hCG in the blood.
              3. Free Alpha Subunit of hCG (hCGα): It goes without saying that this is the alpha subunit all by itself.  It has no biological activity.  That is, it doesn't work and can't keep blood progesterone levels high.
              4. Free Beta Subunit of hCG (hCGβ): This variant of hCG is the free beta subunit without the alpha subunit and it also lacks biological activity.  During pregnancy, the free beta subunit makes up about 1% of all the hCG in the blood.  Also, this variant is sometimes made by certain types of cancers.
              5. Nicked Free Beta Subunit of hCG (hCGβn): This is similar to hCGn (#2 above) yet there is no alpha subunit attached to it and so it, too, has no activity.
              6. Beta Core Fragment of hCG (hCGβcf): This is the final breakdown product of hCG.  It is a small molecule that is made up of the center (core) part of the hCG beta subunit.  Like the other non-intact hCG variants, the beta core fragment is inactive.  However, unlike all of the other hCG forms that are present in both the blood and the urine of pregnant women, the beta core fragment is only present in the urine.  In fact, the urine of pregnant women contains more beta core fragment than any of the other hCG variants.
              7. Hyperglycosylated hCG (hCG-h): Intact hCG normally has carbohydrate (sugar) molecules attached to it.  Hyperglycosylated hCG is like intact hCG except it has even more carbohydrate molecules attached.  This hCG variant is the predominant form of hCG produced during the first few weeks of pregnancy but it is nearly undetectable by the end of the first trimester.

               Here is a figure of the forms of hCG that contain all or part of the beta subunit (adapted from Clin Chem 1997;43:2233-2243):

              HCG_ASCP 2011
              All of these forms of hCG (and probably others that we don't even know about) are produced during pregnancy and can be detected in the blood and urine of pregnant women (hCGβcf is only in the urine).  Future posts will discuss how these different hCG variants make detecting and measuring hCG a bit of a challenge.  Things are not always as straightforward as they seem!