Genetic testing approaches

The Testing Arsenal - From Karyotypes to NGS

• Karyotyping: Visualizes entire chromosomes. The foundational tool for detecting large-scale changes.

  • Target: Aneuploidies (e.g., Trisomy 21, Turner syndrome) and large structural rearrangements (e.g., translocations).

• Fluorescence In Situ Hybridization (FISH): Uses fluorescent probes that bind to specific DNA sequences.

  • Target: Known microdeletions/duplications (e.g., 22q11.2 in DiGeorge, Williams syndrome) and confirming rearrangements.

• Chromosomal Microarray (CMA): High-resolution genome-wide scan for copy number variations (CNVs).

  • Target: Gains (duplications) and losses (deletions) of DNA segments, smaller than what karyotyping can detect.

⭐ First-tier test for non-syndromic developmental delay, intellectual disability, and autism spectrum disorder.

• Next-Generation Sequencing (NGS): Massively parallel sequencing of millions of DNA fragments at once.
  • Target: From single gene variants to the entire genome. Includes gene panels, whole-exome (WES), and whole-genome (WGS) sequencing.

Clinical Clues - Which Test When?

• Karyotyping: Suspected aneuploidy (e.g., Down, Turner, Klinefelter syndrome) or balanced translocation (e.g., recurrent miscarriages).
• FISH (Fluorescence In Situ Hybridization): Known microdeletion/duplication syndromes (e.g., DiGeorge, Williams, Prader-Willi/Angelman).
• Chromosomal Microarray (CMA): First-tier for non-specific presentations like multiple congenital anomalies, unexplained developmental delay/intellectual disability (DD/ID), or autism spectrum disorder (ASD).
• Next-Generation Sequencing (NGS):
  • Gene Panel: For genetically heterogeneous disorders where multiple genes cause a similar phenotype (e.g., Noonan syndrome, hearing loss, epilepsy).
  • Whole Exome/Genome Sequencing (WES/WGS): For complex cases, diagnostic odysseys, or when panel testing is negative.

⭐ Exam Favourite: Chromosomal Microarray (CMA) is the first-tier investigation for a child with unexplained Global Developmental Delay (GDD), Intellectual Disability (ID), or Autism Spectrum Disorder (ASD).

Antenatal Answers - Peeking Before Birth

• Prenatal Screening (Non-invasive): Assesses risk, not a definitive diagnosis.

  • First Trimester Combined Test (11-14 wks):
    • Double Marker: Free β-hCG + PAPP-A.
    • Nuchal Translucency (NT) Scan.
  • Second Trimester Quadruple Test (15-20 wks):
    • AFP, β-hCG, Unconjugated Estriol (uE3), Inhibin A.
  • Non-Invasive Prenatal Testing (NIPT):
    • Analyzes cell-free fetal DNA (cffDNA) in maternal blood from 10 weeks.
    • High accuracy for common aneuploidies.

• Prenatal Diagnosis (Invasive): Confirmatory tests.

  • Chorionic Villus Sampling (CVS):
    • Done at 10-13 weeks; allows earlier diagnosis.
  • Amniocentesis:
    • Done at 15-20 weeks; lower risk of fetal loss than CVS.

⭐ CVS provides an earlier diagnosis than amniocentesis, but with a slightly higher risk of miscarriage.

High‑Yield Points - ⚡ Biggest Takeaways

• Karyotyping is the primary test for aneuploidy (e.g., Trisomy 21) and large structural rearrangements.
• FISH provides rapid detection of specific microdeletions and aneuploidies (e.g., 22q11.2).
• Chromosomal Microarray (CMA) is the first-tier test for unexplained developmental delay & multiple congenital anomalies.
• Sanger sequencing is the gold standard for confirming single-gene mutations found by other methods.
• Next-Generation Sequencing (NGS) is best for genetically heterogeneous disorders.
• MLPA excels at detecting exon-level deletions/duplications (e.g., in DMD).