Basic Introduction to Cytogenetics

Cytogenetics is the branch of genetics that studies the structure and behaviour of chromosomes and their relation to human disease and disease processes.

Cytogenetic studies show the significant contribution of chromosome aberrations to infertility, pregnancy loss at all stages of gestation, and developmental disabilities and congenital malformations in the newborn. The study of human chromosomes also contributed to the understanding of the relation of genetics to the origin of cancers, the physical mapping of genes, and the prevention of genetic diseases by prenatal testing.

Specimen preparation varies with the type of tissue undergoing chromosome analysis. In most instances, the tissues are cultured in a synthetic media to increase the number of cells in mitosis available for cytogenetic analysis. After the culturing, the specimen undergoes a process of harvesting and staining in preparation for microscopic examination of the chromosomes. The time required to complete the chromosomal analysis varies with the type of tissue.

One complete set of human chromosomes consists of 23 chromosomes. A complete set is referred to as the haploid number (N); for humans, N = 23. A normal human somatic cell contains two complete sets of chromosomes, one of maternal origin and the other of paternal origin. There are 46 chromosomes in the nuclei of all human somatic cells; this number is known as the diploid number (2N); for humans, 2N = 46.

Chromosomes are classified as autosomes or as sex chromosomes. There are 22 autosomes in the human chromosome complement. Because a somatic cell contains two members of each chromosome, there are 22 homologous (morphologically similar) pairs in human cells. Cells with two X chromosomes are classified as female, and cells with one X and one Y chromosome are classified as male. These two chromosomes are the sex chromosomes.

A routine cytogenetic analysis involves evaluating cells to determine their modal chromosome number and assessing the structural integrity of each chromosome in the complement. Identification of an extra or missing chromosome is possible by microscopic examination, and the finding is confirmed through the preparation of a set of karyotypes. A karyotype is a photographic or computer-generated representation of the chromosomes in a cell arranged according to size, centromeric position, and banding pattern. Assessment of the structural integrity of each chromosome in the complement can also be made through the microscope, and gross structural chromosome aberrations can be detected as well.

Chromosomal analysis can be performed on cells derived from many sources, including peripheral blood lymphocytes, fetal blood, chorionic villi, amniotic fluid cells, skin and bone marrow.

Neoplastic Cytogenetics

Since the discovery of the Philadelphia chromosome in chronic myelogeneous leukaemia (CML) by Nowell and Hungerford (1960), the clinical uses of cytogenetics as a tool for diagnosis, prognosis, staging, monitoring response to treatment, and understanding human malignancies have expanded dramatically, both for haematological disorders and for many solid tumours.
An example of a cancer cytogenetic karyotype depicting the classic Philadelphia rearrangement; 46,XX,t(9;22)(q34;q11.2)

Culturing of malignant cells for cytogenetic analysis requires a working knowledge of the diseases, tissue types, culture parameters, and harvest strategies. The quality of the chromosomes obtained from malignant cells is often inferior to that of constitutional chromosome studies. It is important to understand that only the malignant cells will exhibit the malignancy-associated clonal cytogenetic abnormalities. If no malignant cells are dividing, the result will often be a normal karyotype derived from normal cells present in the sample.

Bone marrow is the specimen of choice for cancer cytogenetics as it exhibits the abnormal karyotypes more reliably than peripheral blood. Peripheral blood is useful in cases where the diseased cells have migrated out of the bone marrow and are circulating in the peripheral blood.
Blood may also be used in certain cases such as Chronic Lymphoblastic leukaemia and in some lymphomas, or when blasts are present in a high proportion of the white blood cells.

1.5-2ml of sterilely collected lithium/sodium heparinised bone marrow, preferably the first aspirate, as blood dilutes the bone marrow with subsequent aspirates, is sufficient for cytogenetic analysis.

Results are routinely available in 14-21 days.

Samples should be kept cool in transit, protected from extremes in temperature, and reach the laboratory as quickly as possible. The malignant cells present in the sample will only divide a few times before becoming inactive and therefore after three days in transit, a successful culture may become less likely.

Cytogenetic analysis of chromosomes in cancer has been exploited as a powerful diagnostic tool, as a means of gaining prognostic information, and as a means of identifying markers for follow-up studies of the malignant clone. Cancer cytogenetic analysis is now an established routine component of the management of patients with haematologic malignancies, from diagnosis and prognosis, through treatment, remission, relapse, transplant and post-transplant care.

Dunn, B., Mouchrani, P. and Keagle, M. ed., (1994). The Cytogenetic Symposia. 2nd ed. Texas: Association of Genetic Technologists, pp7;1-7. 13;1-5.

Postnatal Cytogenetics

Constitutional chromosome analysis is performed on peripheral blood samples collected in lithium/sodium-heparin tubes. We require 2ml of peripheral blood from adults and 1-2ml of peripheral blood from babies and thereafter metaphases are derived from cultured blood lymphocytes for analysis.

Samples should be kept cool in transit, protected from extremes in temperature, and reach the laboratory as quickly as possible.

The main referral reasons for cytogenetic analyses are:

  • Intellectual disability or developmental delay
  • Infertility
  • Abnormalities of sexual development
  • Recurrent pregnancy loss
  • Dysmorphism
  • Congenital abnormalities.

Referrals are sent via general practitioners or specialist clinics. Reports are available within 14 days for routine samples, or within 10 days for urgent referrals. Urgent samples include neonates, parents of an ongoing pregnancy or specific clinical requests.

Chromosome studies may provide a diagnosis in cases of intellectual disability or congenital abnormalities, or may reveal familial rearrangements responsible for recurrent pregnancy loss. Genetic counselling is offered with an associated genetic counsellor.
Below is an example of a normal female karyotype (46,XX) and a normal male karyotype (46,XY):

Normal female, 46,XX

Normal male, 46,XY

Below is an example of a male karyotype depicting a carrier of a balanced Robertsonian translocation: 45,XY,rob(13;14)(q10)(q10).

This (13;14) translocation is one of the most common Robertsonian translocations observed.

Carriers of Robertsonian translocations are at risk for miscarriages and for offspring with intellectual disability and birth defects associated with aneuploidy, and rarely, uniparental disomy (UPD) which is the inheritance of both copies of a chromosome pair from a single parent. Only those Robertsonian translocation chromosomes containing chromosomes 13 or 21 are associated with an increased risk for having liveborn trisomic offspring.

Gersen, S. and Keagle, M. (2005). The principles of clinical cytogenetics. Totowa, N.J.: Humana Press. Pg 195.

Below is an example of a baby diagnosed with Patau Syndrome (also known as Trisomy 13).

Prenatal Cytogenetics

Parents are always concerned about the health of their unborn baby. Most women give birth to healthy babies, but certain mothers have an increased risk of giving birth to a baby with a birth or genetic defect. Personal and family medical histories provide important information together with physical exams, various procedures (sonography, sonar images) and biochemistry to help identify women who may be at risk to have a baby with a genetic or birth defect. Pregnant women with an increased risk benefit from prenatal diagnostic procedures.

There are three different prenatal diagnostic procedures that may be offered to parents where the pregnancy is at risk, namely amniocentesis (Amnio), chorionic villus sampling (CVS) and cordocentesis (cord blood sampling). The procedure followed is determined by the pregnancy duration and the choice of the consulting doctor (gynaecologist and/or sonographer).


Couples, in consultation with their doctor, can decide on the benefit of the information provided by the results of the prenatal diagnostic procedure.

Each case is different and decisions should be taken after consultation with all parties. Prenatal diagnostic procedures may be recommended for:

  • Women who will be 35 or older at delivery due to an increased risk for a baby with a chromosome abnormality.
  • Couples who have had a baby with a known genetic, chromosome or birth defect.
  • Couples who have a relative with a chromosome abnormality (e.g. Down syndrome), intellectual disability, a medical history of a genetic disorder, or other birth defects.
  • Couples who have experienced two or more pregnancy losses (spontaneous abortions, stillbirths, or neonatal deaths).
  • Couples with an increased risk of having a baby affected with Down syndrome (or other chromosome defect) based on first or second trimester prenatal screening.
  • Couples who are concerned about having an abnormal baby (especially maternal anxiety).


Amniocentesis is a medical procedure in which a small amount of amniotic fluid is withdrawn from the amniotic sac surrounding the fetus in the uterus. An amniocentesis is ideally performed between 14 and 22 weeks gestation.


A sample of approximately 15ml sterilely collected amniotic fluid is sufficient for cytogenetic analysis. Samples should be kept cool in transit, protected from extreme temperatures, and reach the laboratory as soon as possible.

Results are routinely available in 14-21 days.

Chorionic villus sampling, often referred to as CVS, is a diagnostic procedure performed between 10 – 13 weeks gestation. During the test, a small sample of cells (called chorionic villi) are obtained from the placenta more or less where it attaches to the wall of the uterus. This test is to identify chromosome abnormalities and other inherited disorders which may be recommended by your health care provider.


A sample of sterilely collected chorionic villi is collected for cytogenetic analysis. Samples should be kept cool in transit, protected from extreme temperatures, and reach the laboratory as soon as possible.

Results are routinely available in 14-21 days.

Cordocentesis is foetal blood sampling from the umbilical vein and is a diagnostic genetic test that examines blood from the foetal umbilical cord to detect foetal abnormalities. This procedure is performed after 22 weeks gestation, mostly in cases where patients visit the doctor too late for any other prenatal procedure. This foetal blood sample is used for culture and foetal chromosomal analysis and/or foetal DNA extraction for molecular diagnostic procedures.


1.5-2ml of sterilely collected sodium heparinised cord blood is sufficient for cytogenetic analysis. Samples should be kept cool in transit, protected from extreme temperatures, and reach the laboratory as soon as possible.

Results are routinely available in 10-14 days.


In the laboratory, the cells obtained from these prenatal procedures are cultured in the laboratory using special nutrient medium to sustain growth. Chromosomes obtained from the cultured prenatal cells are then analysed. These prenatal tests accurately detect most chromosome abnormalities.


Chromosomes are units of genetic information. Babies with a chromosome abnormality usually have mild to severe intellectual disability and birth defects. One of the most common chromosome abnormalities seen in prenatal testing is Down syndrome. Down syndrome is a genetic disorder caused when abnormal cell division results in an extra copy of chromosome 21.

A normal chromosome complement consists of 46 chromosomes (44 autosomes and two sex chromosomes). A female complement is annotated as 46,XX and a male complement as 46,XY. Cytogenetic tests can reveal the chromosomal sex of the foetus, de novo chromosomal abnormalities, or inherited chromosome abnormalities. Chromosome abnormalities are caused at conception when an abnormal sperm or egg from one parent fuses with a normal sperm or egg from the other parent. The abnormal sperm or egg contains extra or missing chromosome material. These abnormal sperm or eggs appear to be present in everyone, but the risk of an abnormal conception increases significantly with the parents’ ages.

Numeric chromosome abnormalities result from the following:

  • Errors during dividing of sex cells (meiosis)
  • Errors during dividing of somatic cells (mitosis)

Below is an example of a karyotype result obtained from prenatal samples: 47,XY,+21