Chromosome Testing

Chromosome testing can be accomplished using several techniques, all of which can identify chromosomal abnormalities. Karyotyping is the staining, organization, and visualization of chromosomes, which can help identify aneuploidy and major structural changes. Karyotyping is not sensitive in identifying small abnormalities and is a labor-intensive process. Chromosomal microarray analysis is a comparative technique that utilizes fluorescence to identify and quantify specific genetic sequences and is much more sensitive in identifying copy number variants, such as microdeletions or microduplications. However, chromosomal microarray analysis is not useful in identifying certain variations such as balanced translocations. Fluorescence in situ hybridization utilizes fluorescent probes to identify and locate specific genes on chromosomes. Compared with karyotyping, FISH is much more sensitive and specific in determining several abnormalities (except for point mutations) but is limited by the currently available gene probes.

Last updated: 7 Feb, 2022

Editorial responsibility: Stanley Oiseth, Lindsay Jones, Evelin Maza

Overview

Review of the terminology used in medical genetics Genetics Genetics is the study of genes and their functions and behaviors. Basic Terms of Genetics

Numerical chromosomal aberrations: 

  • Aneuploidy: incorrect number of individual chromosomes Chromosomes In a prokaryotic cell or in the nucleus of a eukaryotic cell, a structure consisting of or containing DNA which carries the genetic information essential to the cell. DNA Types and Structure
  • Mosaicism: the presence of ≥ 2 populations of cells within an organism, each with different genomes

Structural chromosomal aberrations:

  • Deletions: the loss of a specific chromosome Chromosome In a prokaryotic cell or in the nucleus of a eukaryotic cell, a structure consisting of or containing DNA which carries the genetic information essential to the cell. Basic Terms of Genetics section
  • Duplications: multiplication of individual chromosome Chromosome In a prokaryotic cell or in the nucleus of a eukaryotic cell, a structure consisting of or containing DNA which carries the genetic information essential to the cell. Basic Terms of Genetics sections
  • Inversions: 180° rotation Rotation Motion of an object in which either one or more points on a line are fixed. It is also the motion of a particle about a fixed point. X-rays of a chromosome Chromosome In a prokaryotic cell or in the nucleus of a eukaryotic cell, a structure consisting of or containing DNA which carries the genetic information essential to the cell. Basic Terms of Genetics section and subsequent reintegration into the original chromosome Chromosome In a prokaryotic cell or in the nucleus of a eukaryotic cell, a structure consisting of or containing DNA which carries the genetic information essential to the cell. Basic Terms of Genetics
  • Translocations:
    • Exchange of fragments between nonhomologous chromosomes Chromosomes In a prokaryotic cell or in the nucleus of a eukaryotic cell, a structure consisting of or containing DNA which carries the genetic information essential to the cell. DNA Types and Structure
    • May be balanced (equal) or unbalanced (unequal)

Cytogenetic testing

Karyotyping

Definition

Karyotyping, also known as chromosomal analysis, is a common method of producing an image of an individual’s chromosomes Chromosomes In a prokaryotic cell or in the nucleus of a eukaryotic cell, a structure consisting of or containing DNA which carries the genetic information essential to the cell. DNA Types and Structure to detect:

  • Large structural changes in chromosomes Chromosomes In a prokaryotic cell or in the nucleus of a eukaryotic cell, a structure consisting of or containing DNA which carries the genetic information essential to the cell. DNA Types and Structure
  • Aneuploidy

Procedure

  • Karyotyping can be performed using various types of samples, including:
    • Blood
    • Bone Bone Bone is a compact type of hardened connective tissue composed of bone cells, membranes, an extracellular mineralized matrix, and central bone marrow. The 2 primary types of bone are compact and spongy. Bones: Structure and Types marrow
    • Amniotic fluid Amniotic fluid A clear, yellowish liquid that envelopes the fetus inside the sac of amnion. In the first trimester, it is likely a transudate of maternal or fetal plasma. In the second trimester, amniotic fluid derives primarily from fetal lung and kidney. Cells or substances in this fluid can be removed for prenatal diagnostic tests (amniocentesis). Placenta, Umbilical Cord, and Amniotic Cavity
    • Placental tissue
  • Method:
    • Cells from a sample are cultured and stimulated to divide.
    • Colchicine Colchicine A major alkaloid from colchicum autumnale l. And found also in other colchicum species. Its primary therapeutic use is in the treatment of gout. Gout Drugs is added to cultured cells → halts cell cycle Cycle The type of signal that ends the inspiratory phase delivered by the ventilator Invasive Mechanical Ventilation in metaphase Metaphase The phase of cell nucleus division following prometaphase, in which the chromosomes line up across the equatorial plane of the spindle apparatus prior to separation. Cell Cycle chromosomes Chromosomes In a prokaryotic cell or in the nucleus of a eukaryotic cell, a structure consisting of or containing DNA which carries the genetic information essential to the cell. DNA Types and Structure are maximally contracted (easier to recognize banding patterns)
    • Chromosomes Chromosomes In a prokaryotic cell or in the nucleus of a eukaryotic cell, a structure consisting of or containing DNA which carries the genetic information essential to the cell. DNA Types and Structure are then:
      • Stained (allows visualization of banding patterns and chromosome Chromosome In a prokaryotic cell or in the nucleus of a eukaryotic cell, a structure consisting of or containing DNA which carries the genetic information essential to the cell. Basic Terms of Genetics identification Identification Defense Mechanisms)
      • Ordered
      • Numbered (23 pairs)
    • Chromosomes Chromosomes In a prokaryotic cell or in the nucleus of a eukaryotic cell, a structure consisting of or containing DNA which carries the genetic information essential to the cell. DNA Types and Structure are analyzed according to:
      • Morphology
      • Size
      • Arm-length ratio
      • Banding patterns
Mitosis

Review of the stages of mitosis Mitosis A type of cell nucleus division by means of which the two daughter nuclei normally receive identical complements of the number of chromosomes of the somatic cells of the species. Cell Cycle: prophase Prophase The first phase of cell nucleus division, in which the chromosomes become visible, the cell nucleus starts to lose its identity, the spindle apparatus appears, and the centrioles migrate toward opposite poles. Cell Cycle, prometaphase Prometaphase The phase of cell nucleus division following prophase, when the breakdown of the nuclear envelope occurs and the mitotic spindle apparatus enters the nuclear region and attaches to the kinetochores. Cell Cycle, metaphase Metaphase The phase of cell nucleus division following prometaphase, in which the chromosomes line up across the equatorial plane of the spindle apparatus prior to separation. Cell Cycle, anaphase Anaphase The phase of cell nucleus division following metaphase, in which the chromatids separate and migrate to opposite poles of the spindle. Cell Cycle, and telophase Telophase The final phase of cell nucleus division following anaphase, in which two daughter nuclei are formed, the cytoplasm completes division, and the chromosomes lose their distinctness and are transformed into chromatin threads. Cell Cycle:
During karyotyping, the cell cycle Cycle The type of signal that ends the inspiratory phase delivered by the ventilator Invasive Mechanical Ventilation is halted in metaphase Metaphase The phase of cell nucleus division following prometaphase, in which the chromosomes line up across the equatorial plane of the spindle apparatus prior to separation. Cell Cycle, usually by adding an agent like colchicine Colchicine A major alkaloid from colchicum autumnale l. And found also in other colchicum species. Its primary therapeutic use is in the treatment of gout. Gout Drugs (inhibits microtubule polymerization to halt mitosis Mitosis A type of cell nucleus division by means of which the two daughter nuclei normally receive identical complements of the number of chromosomes of the somatic cells of the species. Cell Cycle).

Image by Lecturio.

Uses

Karyotyping is useful for identifying and diagnosing:

  • Trisomies and monosomies (e.g., Down syndrome Down syndrome Down syndrome, or trisomy 21, is the most common chromosomal aberration and the most frequent genetic cause of developmental delay. Both boys and girls are affected and have characteristic craniofacial and musculoskeletal features, as well as multiple medical anomalies involving the cardiac, gastrointestinal, ocular, and auditory systems. Down syndrome (Trisomy 21)/ trisomy Trisomy The possession of a third chromosome of any one type in an otherwise diploid cell. Types of Mutations 21)
  • Sex Sex The totality of characteristics of reproductive structure, functions, phenotype, and genotype, differentiating the male from the female organism. Gender Dysphoria chromosome Chromosome In a prokaryotic cell or in the nucleus of a eukaryotic cell, a structure consisting of or containing DNA which carries the genetic information essential to the cell. Basic Terms of Genetics disorders (e.g., Klinefelter syndrome Klinefelter syndrome Klinefelter syndrome is a chromosomal aneuploidy characterized by the presence of 1 or more extra X chromosomes in a male karyotype, most commonly leading to karyotype 47,XXY. Klinefelter syndrome is associated with decreased levels of testosterone and is the most common cause of congenital hypogonadism. Klinefelter Syndrome/47, XXY XXY Klinefelter syndrome is a chromosomal aneuploidy characterized by the presence of 1 or more extra X chromosomes in a male karyotype, most commonly leading to karyotype 47,XXY. Klinefelter syndrome is associated with decreased levels of testosterone and is the most common cause of congenital hypogonadism. Klinefelter Syndrome)
  • Structural abnormalities in chromosomes Chromosomes In a prokaryotic cell or in the nucleus of a eukaryotic cell, a structure consisting of or containing DNA which carries the genetic information essential to the cell. DNA Types and Structure (e.g., Philadelphia chromosome Chromosome In a prokaryotic cell or in the nucleus of a eukaryotic cell, a structure consisting of or containing DNA which carries the genetic information essential to the cell. Basic Terms of Genetics/reciprocal (9;22)(q34;q11) translocation)

Advantages

  • The entire genome Genome The complete genetic complement contained in the DNA of a set of chromosomes in a human. The length of the human genome is about 3 billion base pairs. Basic Terms of Genetics can be visualized at once.
  • Good sensitivity Sensitivity Binary classification measures to assess test results. Sensitivity or recall rate is the proportion of true positives. Blotting Techniques and specificity Specificity Specificity is the probability of correctly determining the absence of a condition. Immunoassays for specific conditions
  • A wide range of sample types can be used.

Disadvantages

  • Can only detect major abnormalities and not small changes
  • Interpretation is operator dependent.
  • Labor-intensive process

Chromosomal Microarray Analysis

Definition

Chromosomal microarray analysis is a high-resolution technique that utilizes fluorescent tags to identify small chromosomal imbalances.

Procedure

  • Can cover the whole genome Genome The complete genetic complement contained in the DNA of a set of chromosomes in a human. The length of the human genome is about 3 billion base pairs. Basic Terms of Genetics or can have specific targets (e.g., for known diseases)
  • A grid of glass or silicon is covered with thousands of specific nucleic acid sequences (probes).
  • Sample and control (or comparative) DNA DNA A deoxyribonucleotide polymer that is the primary genetic material of all cells. Eukaryotic and prokaryotic organisms normally contain DNA in a double-stranded state, yet several important biological processes transiently involve single-stranded regions. DNA, which consists of a polysugar-phosphate backbone possessing projections of purines (adenine and guanine) and pyrimidines (thymine and cytosine), forms a double helix that is held together by hydrogen bonds between these purines and pyrimidines (adenine to thymine and guanine to cytosine). DNA Types and Structure are:
    • Cut into fragments
    • Labeled with different fluorescent tags
    • Added to the slide
  • Hybridized via complementary binding with the probes
  • A scanner detects relative gene Gene A category of nucleic acid sequences that function as units of heredity and which code for the basic instructions for the development, reproduction, and maintenance of organisms. Basic Terms of Genetics expression based on the color and intensity of fluorescence (↑ expression → ↑ intensity).

Uses

  • CMA provides comprehensive genetic testing Genetic Testing Detection of a mutation; genotype; karyotype; or specific alleles associated with genetic traits, heritable diseases, or predisposition to a disease, or that may lead to the disease in descendants. It includes prenatal genetic testing. Myotonic Dystrophies for anomalies not detected during karyotyping:
    • Copy number variants:
      • Microdeletions (e.g., DiGeorge syndrome DiGeorge syndrome DiGeorge syndrome (DGS) is a condition caused by a microdeletion at location q11.2 of chromosome 22 (thus also called 22q11.2 syndrome). There is a defective development of the third and fourth pharyngeal pouches, leading to thymic and parathyroid hypoplasia (causing T-cell immunodeficiency and hypocalcemia, respectively). DiGeorge Syndrome/microdeletion at 22q11.2)
      • Microduplications
      • Unbalanced translocations
    • Single nucleotide polymorphisms
  • Can also detect aneuploidy
  • Used for:

Advantages

  • Can profile the expression of thousands of genes Genes A category of nucleic acid sequences that function as units of heredity and which code for the basic instructions for the development, reproduction, and maintenance of organisms. DNA Types and Structure simultaneously
  • Can detect very small chromosomal abnormalities

Disadvantages

  • Balanced translocations, inversions, and insertions cannot be detected.
  • Mosaicism can affect Affect The feeling-tone accompaniment of an idea or mental representation. It is the most direct psychic derivative of instinct and the psychic representative of the various bodily changes by means of which instincts manifest themselves. Psychiatric Assessment detection.
  • Expensive

Fluorescence In Situ Hybridization

Definition

A technique in which a fluorescent DNA DNA A deoxyribonucleotide polymer that is the primary genetic material of all cells. Eukaryotic and prokaryotic organisms normally contain DNA in a double-stranded state, yet several important biological processes transiently involve single-stranded regions. DNA, which consists of a polysugar-phosphate backbone possessing projections of purines (adenine and guanine) and pyrimidines (thymine and cytosine), forms a double helix that is held together by hydrogen bonds between these purines and pyrimidines (adenine to thymine and guanine to cytosine). DNA Types and Structure (or RNA RNA A polynucleotide consisting essentially of chains with a repeating backbone of phosphate and ribose units to which nitrogenous bases are attached. RNA is unique among biological macromolecules in that it can encode genetic information, serve as an abundant structural component of cells, and also possesses catalytic activity. RNA Types and Structure) probe Probe A device placed on the patient’s body to visualize a target Ultrasound (Sonography) is utilized to identify and locate specific genes Genes A category of nucleic acid sequences that function as units of heredity and which code for the basic instructions for the development, reproduction, and maintenance of organisms. DNA Types and Structure on chromosomes Chromosomes In a prokaryotic cell or in the nucleus of a eukaryotic cell, a structure consisting of or containing DNA which carries the genetic information essential to the cell. DNA Types and Structure

Procedure

  • A single-stranded DNA DNA A deoxyribonucleotide polymer that is the primary genetic material of all cells. Eukaryotic and prokaryotic organisms normally contain DNA in a double-stranded state, yet several important biological processes transiently involve single-stranded regions. DNA, which consists of a polysugar-phosphate backbone possessing projections of purines (adenine and guanine) and pyrimidines (thymine and cytosine), forms a double helix that is held together by hydrogen bonds between these purines and pyrimidines (adenine to thymine and guanine to cytosine). DNA Types and Structure probe Probe A device placed on the patient’s body to visualize a target Ultrasound (Sonography) with a fluorescent dye is used to bind BIND Hyperbilirubinemia of the Newborn to a specific gene Gene A category of nucleic acid sequences that function as units of heredity and which code for the basic instructions for the development, reproduction, and maintenance of organisms. Basic Terms of Genetics site.
  • Probes are hybridized with sample chromosomes Chromosomes In a prokaryotic cell or in the nucleus of a eukaryotic cell, a structure consisting of or containing DNA which carries the genetic information essential to the cell. DNA Types and Structure on a slide.
  • Fluorescence microscopy is used to detect signals.
Chromosomesalufish

Female human karyotype Karyotype The full set of chromosomes presented as a systematized array of metaphase chromosomes from a photomicrograph of a single cell nucleus arranged in pairs in descending order of size and according to the position of the centromere. Congenital Malformations of the Female Reproductive System subjected to FISH with a probe Probe A device placed on the patient’s body to visualize a target Ultrasound (Sonography) to the Alu sequence (green) and counterstained for DNA DNA A deoxyribonucleotide polymer that is the primary genetic material of all cells. Eukaryotic and prokaryotic organisms normally contain DNA in a double-stranded state, yet several important biological processes transiently involve single-stranded regions. DNA, which consists of a polysugar-phosphate backbone possessing projections of purines (adenine and guanine) and pyrimidines (thymine and cytosine), forms a double helix that is held together by hydrogen bonds between these purines and pyrimidines (adenine to thymine and guanine to cytosine). DNA Types and Structure (red)

Image: “ChromosomesAluFish” by Dietzel65. License: CC BY 2.5

Uses

  • FISH can help directly visualize several chromosomal anomalies, including:
    • Aneuploidy
    • Insertions
    • Translocations
    • Microdeletions (e.g., cri du chat syndrome/microdeletions on the short arm Arm The arm, or “upper arm” in common usage, is the region of the upper limb that extends from the shoulder to the elbow joint and connects inferiorly to the forearm through the cubital fossa. It is divided into 2 fascial compartments (anterior and posterior). Arm: Anatomy of chromosome Chromosome In a prokaryotic cell or in the nucleus of a eukaryotic cell, a structure consisting of or containing DNA which carries the genetic information essential to the cell. Basic Terms of Genetics 5)
    • Microduplications
  • To detect genes Genes A category of nucleic acid sequences that function as units of heredity and which code for the basic instructions for the development, reproduction, and maintenance of organisms. DNA Types and Structure associated with malignancy Malignancy Hemothorax (e.g., HER2 HER2 A cell surface protein-tyrosine kinase receptor that is overexpressed in a variety of adenocarcinomas. It has extensive homology to and heterodimerizes with the EGF receptor, the ERBB-3 receptor, and the ERBB-4 receptor. Activation of the erbB-2 receptor occurs through heterodimer formation with a ligand-bound erbB receptor family member. Targeted and Other Nontraditional Antineoplastic Therapy in breast cancer Breast cancer Breast cancer is a disease characterized by malignant transformation of the epithelial cells of the breast. Breast cancer is the most common form of cancer and 2nd most common cause of cancer-related death among women. Breast Cancer)

Advantages

  • Improved sensitivity Sensitivity Binary classification measures to assess test results. Sensitivity or recall rate is the proportion of true positives. Blotting Techniques and specificity Specificity Specificity is the probability of correctly determining the absence of a condition. Immunoassays compared with classic karyotyping
  • Can be used for both dividing and nondividing cells
  • Multiple probes can be used.

Disadvantages

  • Cannot detect point mutations or very small abnormalities
  • Not all chromosomal regions have a probe Probe A device placed on the patient’s body to visualize a target Ultrasound (Sonography) available (at this time).
  • The correct probe Probe A device placed on the patient’s body to visualize a target Ultrasound (Sonography) must be chosen for diagnosis.

References

  1. The New York-Mid-Atlantic Consortium for Genetic and Newborn Screening Services. (2009). Understanding Genetics: A New York, Mid-Atlantic Guide for Patients and Health Professionals. Genetic Alliance; Washington (DC). Retrieved from https://www.ncbi.nlm.nih.gov/pubmed/23304754
  2. Gonzales, P.R., Carroll, A.J., Korf, B.R. (2016). Overview of Clinical Cytogenetics. Curr Protoc Hum Genet. 89, 8.1.1-8.1.13. Retrieved from https://www.ncbi.nlm.nih.gov/pubmed/27037488
  3. Berisha, S.Z., Shetty, S., Prior, T.W., Mitchell, A.L. (2020). Cytogenetic and molecular diagnostic testing associated with prenatal and postnatal birth defects. Birth Defects Res. 112(4), 293-306. Retrieved from https://www.ncbi.nlm.nih.gov/pubmed/32115903
  4. Wan, T.S. (2917). Cancer Cytogenetics: An Introduction. Methods Mol Biol. 1541, 1-10. Retrieved from https://www.ncbi.nlm.nih.gov/pubmed/27910009
  5. Miller, D.T., et al. (2010). Consensus statement: chromosomal microarray is a first-tier clinical diagnostic test for individuals with developmental disabilities or congenital anomalies. Am J Hum Genet. 86(5), 749-64. Retrieved from https://www.ncbi.nlm.nih.gov/pubmed/20466091
  6. Ajitkumar, A., Jamil, R.T., Mathai, J.K. (2020). Cri Du Chat Syndrome. StatPearls. Jan. Retrieved from http://www.ncbi.nlm.nih.gov/books/NBK482460/
  7. Corbitt, H., Morris, S.A., Gravholt, C.H., et al. (2018). TIMP3 and TIMP1 are risk genes for bicuspid aortic valve and aortopathy in Turner syndrome. PLos Genet. 14(10), e1007692. Retrieved from https://journals.plos.org/plosgenetics/article?id=10.1371/journal.pgen.1007692
  8. Silberbach, M., Roos-Hesselink, J.W., Andersen, N.H., et al. (2018). Cardiovascular Health in Turner Syndrome: A Scientific Statement From the American Heart Association. Circ Genom Precis Med. 11(10), e000048. Retrieved from https://www.ahajournals.org/doi/10.1161/HCG.0000000000000048
  9. Schrijver, I., Zehnder, J.L. (2021). Tools for genetics and genomics: Cytogenetics and molecular genetics. In Tirnauer, J.S. (Ed.), UpToDate. Retrieved December 5, 2021, from https://www.uptodate.com/contents/tools-for-genetics-and-genomics-cytogenetics-and-molecular-genetics
  10. Ishida, C., Gupta, V. (2021). Genetics, molecular testing. [online] StatPearls. Retrieved December 5, 2021, from https://www.ncbi.nlm.nih.gov/books/NBK560712/

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