A karyotype is an individual’s complete set of chromosomes, which are photographed and arranged in order, typically captured during the metaphase of cell division. Chromosomes, which carry genetic information, are structures made of DNA and associated proteins. In eukaryotic cells, chromosomes are found in the nucleus, whereas in prokaryotic cells, chromosomes are located in the cytoplasm.

Human karyotype

23 pairs of chromosomes, 22 pairs of autosomes and 1 pair of sex chromosomes, totaling 46 chromosomes in a normal human cell. Genes or genetic information are carried by these chromosomes. Nucleic acids, composed of carbon (C), hydrogen (H), oxygen (O), nitrogen (N) and phosphorus (P), are the building blocks of genes. Three basic components make up DNA, the material that contains genetic information: a sugar, a phosphate group, and a nitrogenous base. Adenine (A), guanine (G), cytosine (C) and thymine (T) are the four possible nitrogenous bases in DNA. Information expressed through gene expression is stored in the structure of these sites.

Creating a karyotype involves several methodical steps to visualize and analyze an individual’s chromosomes. Here is a detailed explanation of the process:

Sample Collection

• • The first step involves collecting a sample, typically blood, but researchers can also use other cell types such as skin cells, bone marrow cells, or amniotic fluid cells (for prenatal testing). In a blood sample, white blood cells are selected for karyotyping because they contain a nucleus with chromosomes, unlike red blood cells, which lack a nucleus.

Cell Culture

1. 1. The collected white blood cells are cultured in a growth medium to stimulate division. Uses the Phytohemagglutinin Method to Stimulate Lymphocytes (White Blood Cells) to Enter the Mitotic Cell Division Phase. The cell culture is maintained at the optimal body temperature of 37°C for white blood cells, allowing 48 to 72 hours to ensure sufficient cell division.

Arresting Cells in Metaphase

To arrest cells in metaphase, where chromosomes are most condensed and visible, chemicals such as colcemid or colchicine are used. The cells are then treated with a hypotonic solution, causing them to swell. This spreads the chromosomes apart, making them easier to distinguish. A hypotonic solution has a lower concentration of solutes compared to the interior of a cell, causing water to move into the cell by osmosis and resulting in cell swelling, which is useful in karyotyping to spread out the chromosomes for easy visualization and analysis.

Fixation

The cells are fixed using a mixture of methanol and acetic acid. This step preserves the structure of the cells and makes them more rigid, ensuring the chromosomes remain in the same stage.

Slide Preparation

The fixed cells are dropped onto a sterile microscope slide, where they burst and spread out, releasing the chromosomes. The slides are allowed to air dry, helping to spread the chromosomes evenly.

Staining

Researchers stain the chromosomes using Giemsa stain, which binds to the DNA and creates a unique banding pattern (G-banding) for each chromosome pair, aiding in their identification. Depending on specific needs, they may also use other staining techniques such as Q-banding (using quinacrine) or R-banding (reverse G-banding).

Microscopy and Photography

The stained slides are examined under a microscope, and high-quality images of the chromosomes are captured using a camera attached to the microscope.

Karyotype Creation

Individual chromosomes from photographs were arranged in pairs according to size, banding pattern, and centromere position, from largest to smallest, either physically or digitally cropped. The sex chromosomes (X and Y) are usually placed last in the system.

Analysis

Researchers perform karyotype analysis to detect chromosomal abnormalities, including structural abnormalities such as extra or missing chromosomes (eg, Down syndrome with an extra chromosome 21), translocations and deletions, and other genetic disorders.

Applications of Karyotyping

1. Diagnostic Tool:
   • Genetic Disorders:
     • Karyotyping is used to determine and diagnose chromosomal abnormalities. It can diagnose the following diseases related to chromosomal abnormalities:
       • Down Syndrome (Trisomy 21):
         • This genetic disorder is caused by the presence of an extra chromosome 21, resulting in trisomy 21.
       • Turner Syndrome (Monosomy X):
         • only one X chromosome in females.
       • Klinefelter Syndrome (XXY):
         • Males with an extra X chromosome.
       • Edwards Syndrome (Trisomy 18):
         • Caused by an extra chromosome 18.
   • Cancers:
     • Many cancers involve chromosomal changes, such as translocations, deletions, or duplications. For example, chronic myelogenous leukemia (CML) is often associated with the Philadelphia chromosome, a translocation between chromosomes 9 and 22. Karyotyping helps diagnose these malignancies and monitor treatment responses.
   • Congenital Anomalies:
     • Karyotyping is used to identify chromosomal abnormalities in newborns with congenital anomalies (birth defects). It can reveal conditions like Patau syndrome (trisomy 13) or structural anomalies such as deletions or duplications that might explain developmental delays or physical malformations.
2. Prenatal Testing:
   • Detecting Chromosomal Abnormalities:
     • Karyotyping is employed in prenatal testing to detect chromosomal abnormalities in fetuses. Techniques like amniocentesis (sampling amniotic fluid) or chorionic villus sampling (CVS) provide fetal cells for karyotype analysis. This can reveal conditions like Down syndrome, Turner syndrome, and other genetic disorders early in pregnancy.
   • Informed Decision-Making:
     • Early detection of chromosomal abnormalities through prenatal karyotyping helps parents and healthcare providers make informed decisions about the pregnancy and prepare for any special care the baby might need after birth.

3. Research:

   • Studying Chromosome Behavior:
     • In genetic and medical research, karyotyping plays a crucial role in understanding the structure and behavior of chromosomes. It facilitates researchers’ understanding of how genetic information is inherited, how chromosome abnormalities arise, and how chromosomes function during cell division.
   • Understanding Genetic Disorders:
     • By examining the karyotypes of individuals with genetic disorders, researchers can determine patterns and mechanisms behind these conditions. This information helps in developing targeted treatments and preventive measures.
   • Cancer Research:
     • Karyotyping aids in investigating chromosomal alterations associated with various types of cancer. Understanding these changes can help develop new treatment approaches and provide insights into the progression and spread of cancer.
   • Evolutionary Studies:
     • Evolutionary biologists use karyotypes to compare chromosomal configurations among different organisms, thereby providing insights into genetic diversity and evolutionary relationships.