Blood Karyotyping
karyotype is the characteristic chromosome complement of a eukaryote species. The preparation and study of karyotypes is part of cytology and, more specifically, cytogenetics. In normal diploid organisms, autosomal chromosomes are present in two identical copies. There may, or may not, be sex chromosomes. Polyploid cells have multiple copies of chromosomes and haploid cells have single copies. The study of whole sets of chromosomes is sometimes known as karyology.

Most (but not all) species have a standard karyotype. The normal human karyotypes contain 22 pairs of autosomal chromosomes and one pair of sex chromosomes. Normal karyotypes for women contain two X chromosomes and are denoted 46,XX; men have both an X and a Y chromosome denoted 46,XY. However, some individuals have other karyotypes with added or missing chromosomes, and in all such cases there are developmental abnormalities as a consequence.

Karyotypes can be used for many purposes. They may be used to study chromosomal aberrations, to study cellular function, to study taxonomic relationships, or to gather information about past evolutionary events.

Karyotype is done to:
• Determine whether the chromosomes of an adult have an abnormality that can be passed on to a child.
• Determine whether a chromosome defect is preventing a woman from becoming pregnant or causing miscarriages.
• Determine whether a chromosome defect is present in a fetus. Karyotyping also may be done to determine whether chromosomal problems may have caused a fetus to be stillborn.
• Determine the cause of a baby's birth defects or disability.
• Help determine the appropriate treatment for some types of cancer.
• Identify the sex of a person by determining the presence of the Y chromosome. This may be done when a newborn's sex is not clear.

Karyotype testing can be done using almost any cell or tissue from the body. A karyotype test usually is done on a blood sample taken from a vein. For testing during pregnancy, it may also be done on a sample of amniotic fluid or the placenta.

Results of a karyotype test are usually available within 1 to 2 weeks. Patients will find the 46 chromosomes that can be grouped as 22 matching pairs and 1 pair of sex chromosomes (XX for a female and XY for a male) in case of normal result. Also the size, shape, and structure are normal for each chromosome. In case of any abnormality, there are more than or less than 46 chromosomes, he shape or size of one or more chromosomes is abnormal and a chromosome pair may be broken or incorrectly separated.

Sometimes a karyotype test is combined with other genetic tests to provide more specific information about genetic problems. If the results of karyotype are abnormal, other family members may be advised to undergo testing. Since the information obtained from a genetic test can have a profound impact on your life, you may want to see a doctor who specializes in genetics (geneticist) or a genetic counselor. This type of counselor is trained to help you understand your risk for having a child with an inherited (genetic) disease, such as sickle cell disease, cystic fibrosis, or hemophilia. A genetic counselor can help you make well-informed decisions.

Chromosomal Breakage Studies
Chromosomal breakage syndromes are a group of genetic disorders that are typically transmitted in an autosomal recessive mode of inheritance. In culture, cells from affected individuals exhibit elevated rates of chromosomal breakage or instability, leading to chromosomal rearrangements. The disorders are characterized by a defect in DNA repair mechanisms or genomic instability, and patients with these disorders show increased predisposition to cancer.

The following specific chromosome breakage syndromes are addressed in separate subsections of this article:
• Ataxia telangiectasia (AT)
• Bloom syndrome (BS)
• Fanconi anemia (FA)
• Xeroderma pigmentosum (XP)

Chromosomal breakage syndromes are relatively rare; most practicing physicians may never see a patient with a chromosomal breakage syndrome. Some of the specific syndromes occur at relatively high rates in certain ethnic groups. Diagnosis is complicated because the symptoms may be varied and complex. These disorders are often lethal.

In case of Ataxia telangiectasia, risk of neoplasia 38%; 85% are leukemias and lymphomas (B-cell). Young children have acute lymphoblastic leukemia of T-cell origin. Older children have T-cell leukemia. Risk for other cancers is increased 4-fold (2- to 3-fold increased risk for breast cancer in carriers).

In Bloom syndrome <1% of mutations, risk for leukemia, lymphoma, adenocarcinoma, and other cancers increased 5- to 8-fold.

Fanconi anemia specially the aplastic anemia increases the risk for pancytopenia, leukemia, acute myeloid leukemia, squamous cell carcinoma, medulloblastoma, Wilms tumor, breast cancer, and liver cancer.

Xeroderma pigmentosum increases the risk of squamous cell carcinoma, basal cell carcinoma, malignant melanoma, and fibrosarcoma by 1000 fold and 10- to 20-fold increased risk for other tumors.

Sister Chromatid Exchange
Sister Chromatid Exchange (SCE) represents the interchange of homologous segments between two chromatids of one chromosome. SCEs can be detected by growing cells under special culture conditions to produce differential staining of sister chromatids. SCE analysis aids in the diagnosis of inherited conditions such as Bloom syndrome, which shows an increased rate of SCE compared to controls. Because special culture conditions are required, chromosome breakage studies must be specifically requested.

The SCE test is usually performed on human peripheral blood lymphocytes.

The advantage of using sister chromatid exchange analysis are
• Cell/ cell approach
• Possible co-detection of cell proliferation rate (ratio M1/M2/M3)
• Assessment of high frequency cells (HFC) with high statistical power

Karyotyping, Amniotic Fluid
Karyotyping is a test to examine chromosomes in a sample of cells, which can help identify genetic problems as the cause of a disorder or disease. This test can:

• Count the number of chromosomes
• Look for structural changes in chromosomes


This test is usually done to evaluate a couple with a history of miscarriages or to examine an abnormal appearance of the body that suggests a genetic abnormality.

The bone marrow or blood test can be done to identify the Philadelphia chromosome, which is found in about 85% of those with chronic myelogenous leukemia (CML).

The amniotic fluid test is done to check a developing fetus for chromosome abnormalities.

Normal Result
• Females: 44 autosomes and 2 sex chromosomes (XX), denoted 46, XX
• Males: 44 autosomes and 2 sex chromosomes (XY), denoted 46, XY

Abnormal Results
Abnormal results may be due to:
• Down syndrome
• Klinefelter syndrome
• Philadelphia chromosome
• Trisomy 18
• Turner syndrome

This list is not all-inclusive.

Additional conditions under which the test may be performed:
• Chronic myelogenous leukemia (CML) or other leukemias
• Multiple birth defects
• Ambiguous genitalia

Amniotic fluid is collected from mother’s amniotic cavity. This is an invasive procedure in which a needle is passed through the mother's lower abdomen into the amniotic cavity inside the uterus. Enough amniotic fluid is present for this to be accomplished starting about 14 weeks gestation. For prenatal diagnosis, most amniocenteses are performed between 14 and 20 weeks gestation. The increased risk for fetal mortality following amniocentesis is about 0.5% above what would normally be expected.

Karyotyping, Amniotic Fluid with AFP
Prenatal diagnosis employs a variety of techniques including the chromosomal analysis through Karyotyping of the amniotic fluid to determine the health and condition of an unborn fetus. Without knowledge gained by prenatal diagnosis, there could be an untoward outcome for the fetus or the mother or both. Congenital anomalies account for 20 to 25% of perinatal deaths. Specifically, prenatal diagnosis is helpful for:
• Managing the remaining weeks of the pregnancy
• Determining the outcome of the pregnancy
• Planning for possible complications with the birth process
• Planning for problems that may occur in the newborn infant
• Deciding whether to continue the pregnancy
• Finding conditions that may affect future pregnancies


There are a variety of non-invasive and invasive techniques available for prenatal diagnosis. Each of them can be applied only during specific time periods during the pregnancy for greatest utility. The techniques employed for prenatal diagnosis include:
• Ultrasonography
• Amniocentesis
• Chorionic villus sampling
• Fetal blood cells in maternal blood
• Maternal serum alpha-fetoprotein
• Maternal serum beta-HCG
• Maternal serum estriol

The developing fetus has two major blood proteins--albumin and alpha-fetoprotein (AFP). Since adults typically have only albumin in their blood, the MSAFP test can be utilized to determine the levels of AFP from the fetus. Ordinarily, only a small amount of AFP gains access to the amniotic fluid and crosses the placenta to mother's blood. However, when there is a neural tube defect in the fetus, from failure of part of the embryologic neural tube to close, then there is a means for escape of more AFP into the amniotic fluid. Neural tube defects include anencephaly (failure of closure at the cranial end of the neural tube) and spina bifida (failure of closure at the caudal end of the neural tube). The incidence of such defects is abbout 1 to 2 births per 1000 in the United States. Also, if there is an omphalocele or gastroschisis (both are defects in the fetal abdominal wall), the AFP from the fetus will end up in maternal blood in higher amounts.

The MSAFP has the greatest sensitivity between 16 and 18 weeks gestation, but can still be useful between 15 and 22 weeks gestation.

However, the MSAFP can be elevated for a variety of reasons which are not related to fetal neural tube or abdominal wall defects, so this test is not 100% specific.

The MSAFP can also be useful in screening for Down syndrome and other trisomies. The MSAFP tends to be lower when Down syndrome or other chromosomal abnormalities is present.

Karyotyping, Bone Marrow
Karyotyping is a test to examine chromosomes in a sample of cells, which can help identify genetic problems as the cause of a disorder or disease.
This test can:
• Count the number of chromosomes
• Look for structural changes in chromosomes

The test can be performed on a sample of blood, bone marrow, amniotic fluid, or tissue from the placenta, the organ that develops during pregnancy to feed a growing baby.

Bone marrow karyotyping is usually required to identify the leukemias. A bone marrow specimen requires a bone marrow biopsy. The sample is placed into a special dish and allowed to grow in the laboratory. Cells are later taken from the growing sample and stained. The laboratory specialist uses a microscope to examine the size, shape, and number of chromosomes in the cell sample. The stained sample is photographed to provide a karyotype, which shows the arrangement of the chromosomes.

Certain abnormalities can be identified through the number or arrangement of the chromosomes. Chromosomes contain thousands of genes that are stored in DNA, the basic genetic material.

The bone marrow or blood test can be done to identify the Philadelphia chromosome, which is found in about 85% of those with chronic myelogenous leukemia (CML).

In some cases, an abnormality may occur as the cells as growing in the lab dish. Karyotype tests should be repeated to confirm that an abnormal chromosome problem is actually in the body of the patient.

Karyotyping, Product of conception
Chromosomal abnormalities such as, an extra or missing chromosome, deletions or additions to the normal structure of specific chromosomes, or translocations between two chromosomes have been identified and correlated with specific phenotypes of anatomical, developmental and physiological abnormalities.
Roughly 10-15% of all patients submitted for such testing by infertility clinics have an abnormal chromosomal picture. While some are acquired by inheritance, the majority originate anew. Chromosome abnormalities are a significant cause of recurrent miscarriage and/or infertility.

After three miscarriages there is a 3-8% chance that one member of the couple carries a balanced chromosome rearrangement (i.e. translocation) that can become unbalanced in reproduction. These translocations may be responsible for either recurrent miscarriage or prolonged infertility. Chromosome analysis is recommended for those couples undergoing infertility evaluation, those with recurrent miscarriage, men with oligo- or azoospermia, women with primary amenorrhea, or those individuals with a family history of a chromosome abnormality or mental retardation.

Chromosome analysis of products of conception (POC) is recommended for recurrent miscarriages as 50% of first trimester miscarriages are due to chromosome abnormalities. This analysis is helpful for the couple to discover the cause of miscarriages, to better calculate recurrence risks for chromosome abnormality, and to possibly preclude further evaluation of other causes of pregnancy loss.

Karyotyping, Fragile X Syndrome
Fragile X is a family of genetic conditions, which can impact individuals and families in various ways. These genetic conditions are related in that they are all caused by gene changes in the same gene, called the FMR1 gene.

Fragile X syndrome (FXS) is the most common cause of inherited mental impairment. This impairment can range from learning disabilities to more severe cognitive or intellectual disabilities. (Sometimes referred to as mental retardation.) FXS is the most common known cause of autism or "autistic-like" behaviors. Symptoms also can include characteristic physical and behavioral features and delays in speech and language development.

Fragile X can be passed on in a family by individuals who have no apparent signs of this genetic condition. In some families a number of family members appear to be affected, whereas in other families a newly diagnosed individual may be the first family member to exhibit symptoms.

The X chromosome of some people is unusually fragile at one tip - seen "hanging by a thread" under a microscope. Most people have 29 "repeats" at this end of their X-chromosome, those with Fragile X have over 700 repeats due to duplications. This syndrome affects 1:1500 males, 1:2500 females.

Fragile X (DNA)
The fragile X DNA test has revolutionized fragile X syndrome diagnosis and accompanying genetic counseling. It has been available since 1991 and provides definitive diagnosis of fragile X syndrome and extremely accurate carrier detection. Reliable for people of any age, it can also be performed prenatally. It has superseded the fragile X cytogenetic test due to its greater reliability and accuracy in diagnosis and its ability to identify unaffected carriers.

Although fragile X syndrome is the most common cause of inherited intellectual impairment, it is underdiagnosed. Awareness of fragile X syndrome and the utility of the fragile X DNA test is growing among non-geneticist physicians but is not yet widespread. Knowledge of this disorder is particularly important in pediatrics, neurology, obstetrics/gynecology and general practice. Wide variability in the clinical presentation of the disorder is one reason the diagnosis may be missed. Although certain physical and behavioral features are often associated with fragile X syndrome, they are not always present. In at least 10% of cases in males, intellectual impairment is the only presenting sign. The classic triad of long face, prominent ears and macroorchidism is present in just 60% of cases. Mental retardation is not a constant, either. Approximately 15% of males with fragile X syndrome have an IQ above 70 (Hagerman et al., 1994) . In cases such as these the possibility of fragile X syndrome may not be considered. Similarly, females with fragile X syndrome may not be correctly diagnosed because symptoms can be subtle.

Many asymptomatic carriers of fragile X syndrome are unaware they are carriers because there may not be a family history of fragile X syndrome, or because a relative with fragile X syndrome may not have been diagnosed. The carrier rate in females is quite high, at approximately 1/300. It is recommended that fragile X carrier testing be offered to all women of reproductive age who have a relative with mental retardation of unknown cause.

Testing for the fragile X mutation is based primarily on measuring the length of the FMR1 gene region containing the CGG repeat stretch and then calculating the CGG repeat number. Analysis of the gene's methylation status (ie. whether the gene is turned 'off ' or 'on') is often performed simultaneously. Categorization of the mutation type is based on CGG repeat number and in some cases also on the methylation status of the gene. Methylation information is useful for delineating premutations from full mutations when the repeat number is intermediate (~150-250) and can have prognostic value when a full mutation is methylated in only a small percentage of cells.

Chromosome Philadelphia
Philadelphia chromosome or Philadelphia translocation is a specific chromosomal abnormality that is associated with chronic myelogenous leukemia (CML). It is due to a reciprocal translocation which means an exchange of genetic material between region q34 of chromosome 9 and region q11 of chromosome 22. The presence of this translocation is a highly sensitive test for CML, since 95% of people with CML have this abnormality (The remainder have either a cryptic translocation that is invisible on G-banded chromosome preparations, or a variant translocation involving another chromosome or chromosomes as well as the long arm of chromosomes 9 and 22). However, the presence of the Philadelphia (Ph) chromosome is not sufficient to diagnose CML, since it is also found in acute lymphoblastic leukemia (ALL, 25–30% in adult and 2–10% in pediatric cases) and occasionally in acute myelogenous leukemia (AML).

The exact chromosomal defect in Philadelphia chromosome is translocation. Parts of two chromosomes, 9 and 22, swap places. The result is that part of the BCR ("breakpoint cluster region") gene from chromosome 22 (region q11) is fused with part of the ABL gene on chromosome 9 (region q34). The fused "bcr-abl" gene is located on the resulting, shorter chromosome 22. Because abl carries a domain that can add phosphate groups to tyrosine residues (tyrosine kinase) the bcr-abl fusion gene is also a tyrosine kinase. The fused bcr-abl protein interacts with the interleukin 3beta(c) receptor subunit. The bcr-abl transcript is constitutively active, i.e. it does not require activation by other cellular messaging proteins. In turn, bcr-abl activates a number of cell cycle-controlling proteins and enzymes, speeding up cell division. Moreover, it inhibits DNA repair, causing genomic instability and potentially causing the feared blast crisis in CML.

Comprehensive Metabolic Screening

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DMD, DNA Testing
This is an X linked recessive disease. DMD affects only males, with rare exceptions. Unless a boy with DMD is known to be at risk because of his family history, he is unlikely to be diagnosed before the age of 2 or 3 years. Most boys with DMD walk alone at a later age than average. Then the parents are likely to be worried about something unusual in the way he walks, about frequent falling or about difficulty rising from the ground or difficulty going up steps. Less often, concern arises because of intellectual handicap ("mental retardation"). Although intellectual handicap affects only a minority of boys with DMD, it is more frequent than in other children.

DMD, the largest known human gene, provides instructions for making a protein called dystrophin. Duchenne and Becker muscular dystrophy - caused by mutations in the DMD gene. Hundreds of mutations in the DMD gene have been identified in people with the Duchenne and Becker forms of muscular dystrophy. Most of these mutations delete part of the DMD gene. Other mutations abnormally duplicate part of the gene or change a small number of DNA building blocks (nucleotides) in the gene.

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