Alkali denaturation test

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Alkali denaturation test
Purposedifferentiate neonatal from maternal blood

The alkali denaturation test, also known as A or Apt test, is a medical test used to differentiate fetal or neonatal blood from maternal blood found in a newborn's stool or vomit, or from maternal vaginal blood. [1] [2]

Contents

History

The test was developed by Leonard Apt (1922–2013), [3] an American pediatric ophthalmologist. The test was originally used to identify the source of bloody stools in newborn infants. It has been modified to distinguish fetal from maternal hemoglobin in blood samples from any source. [4]

Uses

The Apt test is most commonly used in cases of vaginal bleeding late during pregnancy (antepartum haemorrhage) to determine if the bleeding is from the mother or the fetus. [ citation needed ]

In practice, the Apt test may not be done when there is suspicion of vasa previa, because the time to fetal collapse with bleeding from vasa previa is often very short.

The Apt test can also be used to detect the presence of fetal blood in the maternal circulation in cases of suspected fetal-maternal hemorrhage. Since the test is only a qualitative determination of the presence of fetal hemoglobin in maternal blood, the quantitative Kleihauer-Betke test is more commonly used.

Finally, the Apt test can be used after birth (postpartum hemorrhage) if the newborn has bloody vomiting, bloody stool, or active bleeding from the nasogastric tube. A positive apt test would mean that the blood is either due to gastrointestinal or pulmonary bleeding from the neonate. A negative Apt test would indicate that the blood is of maternal origin, suggesting that the neonate swallowed or aspirated maternal blood, either during delivery or during breastfeeding (e.g., from breast fissures).

Theory

The test is based on differences between maternal and fetal hemoglobin. Maternal blood contains adult hemoglobin composed of two alpha and two beta subunits (aka hemoglobin A or HbA; i.e., normal adult hemoglobin). Fetal blood contains fetal hemoglobin composed of two alpha and two gamma subunits (aka hemoglobin F or HbF; i.e., normal fetal hemoglobin). This difference in composition gives the different types of hemoglobin different chemical properties (in addition to the higher affinity HbF has for dissolved blood oxygen over HbA, allowing baby to extract oxygen from the mother's blood). Fetal hemoglobin is resistant to alkali (basic) denaturation, whereas adult hemoglobin is susceptible to such denaturation. Therefore, exposing the blood specimen to sodium hydroxide (NaOH) will denature the adult but not the fetal hemoglobin. The fetal hemoglobin will appear as a pinkish color under the microscope while the adult hemoglobin will appear as a yellow-brownish color.

Method

The blood is mixed with a small amount of sterile water to cause hemolysis of the RBCs, yielding free hemoglobin. The sample is next centrifuged for several minutes. The pink hemoglobin-containing supernatant is then mixed with 1 mL of 1% NaOH for each 5 mL of supernatant. The color of the fluid is assessed after 2 minutes. Fetal hemoglobin will stay pink and adult hemoglobin will turn yellow-brown since adult hemoglobin is less stable and will convert to hematin which has a hydroxide ligand. [5]

See also

Related Research Articles

Rh disease is a type of hemolytic disease of the fetus and newborn (HDFN). HDFN due to anti-D antibodies is the proper and currently used name for this disease as the Rh blood group system actually has more than 50 antigens and not only the D-antigen. The term "Rh Disease" is commonly used to refer to HDFN due to anti-D antibodies, and prior to the discovery of anti-Rho(D) immune globulin, it was the most common type of HDFN. The disease ranges from mild to severe, and occurs in the second or subsequent pregnancies of Rh-D negative women when the biologic father is Rh-D positive.

<span class="mw-page-title-main">Hematochezia</span> Medical condition

Haematochezia is the passage of fresh blood through the anus, usually in or with stools. The term is from Greek αἷμα ("blood") and χέζειν. Hematochezia is commonly associated with lower gastrointestinal bleeding, but may also occur from a brisk upper gastrointestinal bleed. The difference between hematochezia and rectorrhagia is that, in the latter, rectal bleeding is not associated with defecation; instead, it is associated with expulsion of fresh bright red blood without stools. The phrase bright red blood per rectum is associated with hematochezia and rectorrhagia.

<span class="mw-page-title-main">Fetal hemoglobin</span> Oxygen carrier protein in the human fetus

Fetal hemoglobin, or foetal haemoglobin is the main oxygen carrier protein in the human fetus. Hemoglobin F is found in fetal red blood cells, and is involved in transporting oxygen from the mother's bloodstream to organs and tissues in the fetus. It is produced at around 6 weeks of pregnancy and the levels remain high after birth until the baby is roughly 2–4 months old. Hemoglobin F has a different composition than adult forms of hemoglobin, allowing it to bind oxygen more strongly; this in turn enables the developing fetus to retrieve oxygen from the mother's bloodstream, which occurs through the placenta found in the mother's uterus.

<span class="mw-page-title-main">Placenta praevia</span> Medical condition

Placenta praevia is when the placenta attaches inside the uterus but in a position near or over the cervical opening. Symptoms include vaginal bleeding in the second half of pregnancy. The bleeding is bright red and tends not to be associated with pain. Complications may include placenta accreta, dangerously low blood pressure, or bleeding after delivery. Complications for the baby may include fetal growth restriction.

Antepartum bleeding, also known as antepartum haemorrhage (APH) or prepartum hemorrhage, is genital bleeding during pregnancy after the 28th week of pregnancy up to delivery.

Bloody show or show is the passage of a small amount of blood or blood-tinged mucus through the vagina near the end of pregnancy. It is caused by thinning and dilation of the cervix, leading to detachment of the cervical mucus plug that seals the cervix during pregnancy and tearing of small cervical blood vessels, and is one of the signs that labor may be imminent. The bloody show may be expelled from the vagina in pieces or altogether and often appears as a jelly-like piece of mucus stained with blood. Although the bloody show may be alarming at first, it is not a concern of patient health after 37 weeks gestation.

Obstetrical bleeding is bleeding in pregnancy that occurs before, during, or after childbirth. Bleeding before childbirth is that which occurs after 24 weeks of pregnancy. Bleeding may be vaginal or less commonly into the abdominal cavity. Bleeding which occurs before 24 weeks is known as early pregnancy bleeding.

<span class="mw-page-title-main">Neonatal jaundice</span> Medical condition

Neonatal jaundice is a yellowish discoloration of the white part of the eyes and skin in a newborn baby due to high bilirubin levels. Other symptoms may include excess sleepiness or poor feeding. Complications may include seizures, cerebral palsy, or kernicterus.

<span class="mw-page-title-main">Blood in stool</span> Medical condition

Blood in stool or rectal bleeding looks different depending on how early it enters the digestive tract—and thus how much digestive action it has been exposed to—and how much there is. The term can refer either to melena, with a black appearance, typically originating from upper gastrointestinal bleeding; or to hematochezia, with a red color, typically originating from lower gastrointestinal bleeding. Evaluation of the blood found in stool depends on its characteristics, in terms of color, quantity and other features, which can point to its source, however, more serious conditions can present with a mixed picture, or with the form of bleeding that is found in another section of the tract. The term "blood in stool" is usually only used to describe visible blood, and not fecal occult blood, which is found only after physical examination and chemical laboratory testing.

<span class="mw-page-title-main">Fetal circulation</span> Circulatory system of fetuses

In humans, the circulatory system is different before and after birth. The fetal circulation is composed of the placenta, umbilical blood vessels encapsulated by the umbilical cord, heart and systemic blood vessels. A major difference between the fetal circulation and postnatal circulation is that the lungs are not used during the fetal stage resulting in the presence of shunts to move oxygenated blood and nutrients from the placenta to the fetal tissue. At birth, the start of breathing and the severance of the umbilical cord prompt various changes that quickly transform fetal circulation into postnatal circulation.

<span class="mw-page-title-main">Kleihauer–Betke test</span> Blood test used to measure the amount of fetal hemoglobin

The Kleihauer–Betke ("KB") test, Kleihauer–Betke ("KB") stain, Kleihauer test or acid elution test is a blood test used to measure the amount of fetal hemoglobin transferred from a fetus to a mother's bloodstream. It is usually performed on Rh-negative mothers to determine the required dose of Rho(D) immune globulin (RhIg) to inhibit formation of Rh antibodies in the mother and prevent Rh disease in future Rh-positive children. It is named after Enno Kleihauer and Klaus Betke who described it in 1957.

Rho(D) immune globulin (RhIG) is a medication used to prevent RhD isoimmunization in mothers who are RhD negative and to treat idiopathic thrombocytopenic purpura (ITP) in people who are Rh positive. It is often given both during and following pregnancy. It may also be used when RhD-negative people are given RhD-positive blood. It is given by injection into muscle or a vein. A single dose lasts 12 weeks. It is made from human blood plasma.

<span class="mw-page-title-main">Chignon (medical term)</span> Medical condition

A chignon is a temporary swelling caused by a build-up of bloody fluid left on an infant's head after they have been delivered by vacuum extraction. A vacuum extraction is a type of assistance used during vaginal delivery by an obstetrician or midwife when the second stage of labor, where the cervix is fully dilated allowing for fetus delivery, is stalled. It anatomically resembles regular caput succedaneum, one of two most frequently occurring birth injuries to the head, the other being cephalohematoma, a usually harmless condition where blood accumulates under the newborn's scalp after vaginal delivery.

In ABO hemolytic disease of the newborn maternal IgG antibodies with specificity for the ABO blood group system pass through the placenta to the fetal circulation where they can cause hemolysis of fetal red blood cells which can lead to fetal anemia and HDN. In contrast to Rh disease, about half of the cases of ABO HDN occur in a firstborn baby and ABO HDN does not become more severe after further pregnancies.

<span class="mw-page-title-main">Vasa praevia</span> Condition in which fetal blood vessels cross or run near the internal opening of the uterus

Vasa praevia is a condition in which fetal blood vessels cross or run near the internal opening of the uterus. These vessels are at risk of rupture when the supporting membranes rupture, as they are unsupported by the umbilical cord or placental tissue.

<span class="mw-page-title-main">Percutaneous umbilical cord blood sampling</span>

Percutaneous umbilical cord blood sampling (PUBS), also called cordocentesis, fetal blood sampling, or umbilical vein sampling is a diagnostic genetic test that examines blood from the fetal umbilical cord to detect fetal abnormalities. Fetal and maternal blood supply are typically connected in utero with one vein and two arteries to the fetus. The umbilical vein is responsible for delivering oxygen rich blood to the fetus from the mother; the umbilical arteries are responsible for removing oxygen poor blood from the fetus. This allows for the fetus’ tissues to properly perfuse. PUBS provides a means of rapid chromosome analysis and is useful when information cannot be obtained through amniocentesis, chorionic villus sampling, or ultrasound ; this test carries a significant risk of complication and is typically reserved for pregnancies determined to be at high risk for genetic defect. It has been used with mothers with immune thrombocytopenic purpura.

<span class="mw-page-title-main">Velamentous cord insertion</span> Velamentous placenta

Velamentous cord insertion is a complication of pregnancy where the umbilical cord is inserted in the fetal membranes. It is a major cause of antepartum hemorrhage that leads to loss of fetal blood and associated with high perinatal mortality. In normal pregnancies, the umbilical cord inserts into the middle of the placental mass and is completely encased by the amniotic sac. The vessels are hence normally protected by Wharton's jelly, which prevents rupture during pregnancy and labor. In velamentous cord insertion, the vessels of the umbilical cord are improperly inserted in the chorioamniotic membrane, and hence the vessels traverse between the amnion and the chorion towards the placenta. Without Wharton's jelly protecting the vessels, the exposed vessels are susceptible to compression and rupture.

Fetal-maternal haemorrhage is the loss of fetal blood cells into the maternal circulation. It takes place in normal pregnancies as well as when there are obstetric or trauma related complications to pregnancy.

Early pregnancy bleeding refers to vaginal bleeding before 14 weeks of gestational age. If the bleeding is significant, hemorrhagic shock may occur. Concern for shock is increased in those who have loss of consciousness, chest pain, shortness of breath, or shoulder pain.

<span class="mw-page-title-main">Neonatal infection</span> Human disease

Neonatal infections are infections of the neonate (newborn) acquired during prenatal development or within the first four weeks of life. Neonatal infections may be contracted by mother to child transmission, in the birth canal during childbirth, or after birth. Neonatal infections may present soon after delivery, or take several weeks to show symptoms. Some neonatal infections such as HIV, hepatitis B, and malaria do not become apparent until much later. Signs and symptoms of infection may include respiratory distress, temperature instability, irritability, poor feeding, failure to thrive, persistent crying and skin rashes.

References

  1. Moustafa MH, Taylor M, Fletcher L (August 2005). "My two-week-old daughter is throwing up blood". Acad Emerg Med. 12 (8): 775–7. doi:10.1197/j.aem.2005.03.524. PMID   16079432.
  2. APT L, DOWNEY WS (July 1955). "Melena neonatorum: the swallowed blood syndrome; a simple test for the differentiation of adult and fetal hemoglobin in bloody stools". J. Pediatr. 47 (1): 6–12. doi:10.1016/S0022-3476(55)80118-7. PMID   14392548.
  3. Elaine Schmidt, Obituary: Leonard Apt, 90, doctor-scientist who gave gift of vision to millions of children, UCLA Newsroom, February 7, 2013
  4. Venes, D. Taber's Cyclopedic Medical Dictionary - 20th Ed. F. A. Davis Company, Philadelphia, PA. 2005.
  5. Pediatrics, Gastrointestinal Bleeding

Further reading