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Cytogenetics

Prevent Cross-Contamination

While microbial contamination destroys a culture, cross-contamination destroys the diagnosis. Cross-contamination occurs when cells from “Patient A” are accidentally introduced into the culture vessel of “Patient B.” In Cytogenetics, this is a catastrophic event because the contaminating cells (which are often vigorous, stimulated lymphocytes or fast-growing cell lines) will grow alongside the patient’s cells. This results in a “chimeric” karyotype or, worse, a complete replacement of the patient’s cells, leading to a false report (e.g., reporting a female karyotype for a male patient, or a normal karyotype for a leukemic patient). Preventing this requires strict adherence to workflow discipline

The Golden Rule: One Patient at a Time

The most effective barrier against cross-contamination is the physical and temporal separation of specimens

  • Single-Specimen Workflow: Only one patient’s specimen and their corresponding culture tubes should be inside the Biological Safety Cabinet (BSC) at any given moment
    • Process: Bring Patient A’s rack into the hood \(\rightarrow\) Setup Culture \(\rightarrow\) Close all tubes \(\rightarrow\) Remove Patient A’s rack \(\rightarrow\) Wipe down surface \(\rightarrow\) Bring in Patient B
    • Prohibited: Having multiple open tubes from different patients on the rack simultaneously. This invites error if a pipette tip is reused or accidentally hovered over the wrong tube

Pipetting Discipline

Liquid handling is the primary vector for cell transfer

  • Tip Changes: A pipette tip must never be used for more than one vessel. Even if transferring the same media, if the tip touches the lip of Patient A’s tube and then enters the media bottle, the media bottle is now contaminated with Patient A’s cells. The next patient fed from that bottle will be cross-contaminated
  • Aliquoting Media: Never pour or pipette directly from the stock media bottle into a patient tube if that bottle will be used for others. Instead, pour the required volume of media into a sterile, disposable secondary container (e.g., a 50 mL tube) designated for that specific patient. Discard any excess left in the secondary container
  • Splash Prevention: Avoid vigorous mixing or “blowing out” the pipette. Micro-droplets of cell suspension can splash onto the caps or rims of adjacent tubes or onto the work surface, waiting to be picked up by the next rack

Labeling & Batching Strategy

  • Batching by Sex: When possible, some laboratories batch setups by sex (e.g., set up all Males, then all Females). If a cross-contamination occurs (e.g., a male cell enters a female culture), it is immediately obvious during analysis (finding a Y chromosome in a female case). If two males are cross-contaminated, the error is silent and potentially undetectable
  • Tube positioning: Set up tubes in a consistent order (e.g., Patient Tube on the left, Culture Tube on the right). Never cross hands or reach over open tubes

Maternal Cell Contamination (MCC) Management

A specific form of cross-contamination inherent to prenatal samples (Amnio/CVS) is the contamination of fetal cells with the mother’s cells

  • Dissection (CVS): The laboratory scientist must meticulously remove maternal decidua from fetal villi under a stereomicroscope before culture. This is a manual separation of two different genomes
  • The “First ML” Rule (Amnio): The first 1–2 mL of amniotic fluid drawn is often discarded or used for chemistry because it is most likely to contain maternal blood from the needle stick. Cytogenetics uses the subsequent clean fluid
  • Colony Selection (In Situ): During analysis, laboratory scientists avoid colonies that look morphologically distinct (e.g., epithelial vs. fibroblast) or are growing on the very edge of the coverslip, as these might represent maternal contaminant cells

Sentinel Checks

  • Polymorphisms: Experienced cytogeneticists look for benign chromosomal polymorphisms (e.g., bright satellites on chromosome 21, heterochromatin length on chromosome 9). If a patient’s culture shows two different populations of polymorphisms (e.g., one cell has a large 9qh+ and the next cell does not), cross-contamination is the prime suspect