Scientist in Cytogenetics — CG(ASCP) Preparation Guide Study Guide

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Two blood tubes arrive with matching surnames but different medical record numbers on their orders. Hold the affected accession, preserve specimen integrity and resolve identification through the laboratory's documented discrepancy process before establishing cultures.

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Compare three requests. For a constitutional blood karyotype, collect blood in a culture-compatible heparin container and establish a PHA-stimulated suspension to examine responsive lymphocytes. For a suspected myeloid neoplasm, prioritize a representative marrow aspirate collected with a culture-compatible anticoagulant and promptly establish short-term unstimulated suspensions; a blood lymphocyte culture could miss the target clone. For a skin fibroblast study, submit fresh tissue in a sterile container with suitable transport medium and establish adherent outgrowth; tissue placed entirely in formalin cannot supply viable fibroblasts. If tissue also needs a validated FFPE FISH assay, allocate separate material before fixation. These choices share identity and viability requirements but do not share one stimulation schedule or culture system.

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A delayed marrow specimen produces few metaphases despite adequate total cellularity. Review transport history, viability and culture conditions, and consider complementary testing of the target population before interpreting the limited normal metaphases as exclusionary.

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A chorionic villus specimen contains recognizable villi mixed with decidua. Separate the villi before culture or extraction and retain appropriate material for contamination assessment when indicated, especially if subsequent findings could plausibly reflect maternal cells.

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An unusual chromosome appears repeatedly in one prenatal culture but is absent from another independently established culture. Review colony relationships and specimen-specific mosaicism criteria before deciding whether the observation supports true mosaicism or a culture-associated finding.

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A constitutional blood culture has very few metaphases after an omitted supplement is discovered. If the omitted reagent was phytohaemagglutinin, investigate failure of lymphocyte stimulation rather than first changing the chromosome banding procedure.

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Two cultures handled consecutively develop unexpected, unusually similar findings. Review specimen separation, shared reagents and processing records while evaluating biological explanations. Preserve the original materials so an independent preparation can help distinguish contamination from a real result.

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Two fibroblast flasks reach the planned harvest day, but one remains sparse and the other is densely confluent. Assess each flask's growth and morphology individually before applying the same harvest decision to both cultures.

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A preparation contains many metaphases, but chromosomes are consistently short and highly contracted. Review mitotic inhibitor exposure as a possible contributor before increasing arrest time merely to obtain an even higher metaphase count.

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Metaphases remain tightly clustered despite adequate mitotic activity. Review hypotonic solution preparation, temperature and exposure alongside slide conditions. Adjust one validated parameter at a time so improved spreading can be linked to a specific change.

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A harvest has adequate metaphases but persistent cytoplasmic background. Review fixative preparation and changes, pellet resuspension and handling before altering lymphocyte stimulation, because the observed defect arises after cells have already reached mitosis.

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After a seasonal humidity change, several specimens develop compact spreads despite previously reliable harvests. Review room conditions and slide drying behavior across the batch before changing each specimen's culture or assuming a shared biological abnormality.

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A slide shows dark chromosomes with little internal contrast, while an acceptable control slide bands normally. Review the specimen slide's digestion and preparation characteristics before increasing image contrast or treating the pattern as an unusual chromosome variant.

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Three unrelated cultures fail after introduction of a new medium lot, while cultures established earlier grow normally. Investigate the shared lot and preparation records, preserve evidence and assess other affected cases before attributing each failure to specimen quality.

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A marrow has few metaphases, but the clinical question concerns a specific rearrangement detectable by a validated interphase assay. Use targeted interphase analysis while recognizing that it will not provide a complete conventional karyotype.

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A validated break-apart assay shows a reproducible split pattern above its cutoff. Report the supported rearrangement finding, then use a partner-specific or molecular method if identifying the fusion partner is necessary for the clinical question.

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An IGH/MYC dual-fusion assay yields one separate red signal, one separate green signal and two fusion signals in an abnormal population. Compare this with the assay's validated expected pattern and controls before documenting the rearrangement.

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Tumor nuclei show many clustered target signals while the control usually remains at two copies. This favors disproportionate target gain over simple whole-chromosome polysomy, provided morphology, assay validation and control performance support interpretable nuclei.

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Both target and control signals are weak across a preparation with otherwise recognizable nuclei. Review denaturation, pretreatment and hybridization conditions before interpreting widespread signal absence as simultaneous deletion of multiple unrelated genomic regions.

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A batch develops high diffuse fluorescence after a wash solution preparation error. Recheck composition and temperature against the assay instructions, then evaluate a controlled repeat before altering positivity thresholds to accommodate the background.

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Scattered nuclei in an FFPE section show one signal instead of two, including nuclei in non-neoplastic tissue. Assess truncation background and the validated cutoff before interpreting the tumor as having a clonal deletion.

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Hypothetical example: validation of a particular FFPE assay, using 200 suitable nuclei per specimen, establishes an upper background cutoff of 5.0% for its defined abnormal pattern and a decision rule requiring more than 5.0%. A patient result of 8/200 is 4.0% and does not exceed that cutoff; 24/200 is 12.0% and exceeds it if controls and signal quality are acceptable. A result of 11/200 is 5.5%, only one nucleus above 10/200 at the cutoff. Assess independent-reader agreement, eligibility of the scored nuclei, signal quality and related histologic or laboratory findings. Additional scoring or repeat testing should follow a predefined approach rather than continue until the preferred classification appears. These numbers illustrate interpretation and are not recommended universal thresholds.

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A marrow has a small plasma cell fraction and a negative unselected FISH result. Consider a validated plasma cell enrichment or identification approach before concluding that the clinically relevant plasma cell population lacks the targeted abnormality.

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A patient has a normal copy-number array, but family history suggests a balanced reciprocal translocation. Conventional chromosome analysis can address chromosome structure and arrangement that a dosage-only result does not adequately evaluate.

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An array shows a long homozygous segment on one chromosome with normal copy number. Evaluate possible uniparental inheritance and its clinical relevance rather than reporting the segment as a deletion solely because heterozygous calls are absent.

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A small deletion includes a well-established dosage-sensitive region, while a much larger gain contains limited relevant evidence. Evaluate each through its actual evidence rather than ranking pathogenicity by size or gene count alone.

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A clear metaphase contains 46 replicated structures, each with two visible sister chromatids. Record 46 chromosomes, then assess their identity and structure; the visible pairing does not make the cell a 92-chromosome tetraploid cell.

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A colleague places q21 distal to q31 on a chromosome diagram. Correct the order: increasing band designations move outward from the centromere, so q31 is more distal than q21 on that arm.

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An indication raises concern for a small deletion, but only highly contracted metaphases are available. Improve the preparation or use an appropriate complementary assay rather than interpreting a normal low-resolution karyotype as excluding the suspected lesion.

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Suppose software places image X beside reference A and image Y beside reference B. X instead has B's two p-arm dark bands and q-arm sequence wide L–broad D–L–narrow D. Y has A's single broad p-arm dark band and q-arm sequence narrow D–wide L–D–L–D. Swap X and Y: each then matches its homolog's landmarks in the same centromere-to-telomere order. Return to the source metaphase and trace each segmented chromosome from pter through C to qter, confirming that its bands belong to one continuous chromosome and that no overlap, omitted segment or duplicated image produced the apparent match. If the source shows a genuine unmatched segment, investigate a structural change rather than forcing placement.

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A child's metaphases contain three separate chromosome 21 homologs and a total of 47 chromosomes with XX. Describe the finding as 47,XX,+21 rather than assigning a Robertsonian translocation that was not observed.

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A metaphase has 46 chromosomes, but one homolog has a clear terminal deletion. Classify the finding as structural imbalance rather than rejecting it because the total count remains within the ordinary diploid number.

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A well-preserved metaphase contains three representatives of essentially every chromosome and totals 69. This supports triploidy, whereas a near-diploid cell with several unrelated gains represents an aneuploid pattern rather than three complete sets.

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An adult with recurrent pregnancy loss has an apparently balanced reciprocal translocation and no major health concerns. Explain that normal health does not eliminate the possibility of unbalanced reproductive products, and interpret relevant family studies accordingly.

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A child has 46 chromosomes, including a Robertsonian 14;21 chromosome and two separate chromosome 21 homologs. Recognize three copies of 21q material and investigate parental chromosomes to clarify whether the rearrangement was inherited.

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Band order is reversed between a breakpoint on 3p and another on 3q. Because the inverted segment crosses the centromere, identify the inversion as pericentric before considering its specific breakpoint and inheritance implications.

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A homolog appears longer because a band sequence is repeated, while its partner has the usual pattern. Consider a duplication and use suitable confirmation or genomic characterization to define the gained interval rather than assigning whole-chromosome trisomy.

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A ring chromosome appears in several cells, with other cells showing loss or altered ring forms. Evaluate ring identity and copy-number content while considering mitotic instability as a contributor to the observed cellular variation.

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One X chromosome has two long arms arranged around its centromeric region and lacks the usual short arm. Recognize an isochromosome Xq pattern, then assess the resulting Xp deficiency and Xq dosage in the full complement.

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An unusual chromosome has two centromeric regions and variable secondary structural forms across cells. Consider dicentric-associated instability, but use suitable evidence before claiming that both centromeres are active or that bridging caused every observed change.

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Clinical findings remain suggestive of sex chromosome mosaicism despite an unremarkable blood analysis. Consider whether a clinically relevant second tissue and an appropriately sensitive method could answer the unresolved question rather than assuming uniformity across tissues.

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In cultured amniotic fluid, investigate a single abnormal colony through additional independent colonies or cultures and, when informative, targeted testing of uncultured amniocytes. In CVS, assess direct and cultured findings to characterize the sampled placental populations. If mosaicism is confirmed in villi, follow-up amniocentesis can assess fetal involvement; repeating villus cultures alone cannot answer that question.

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Two marrow metaphases share the same additional chromosome, while two other metaphases each lack a different chromosome. The repeated gain supports a clone; the unrelated single-cell losses do not combine to meet a recurrent-loss criterion.

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A marrow's best-looking metaphases are normal, while less tidy but interpretable cells repeatedly show a shared rearrangement. Include appropriate abnormal cells in the analysis instead of excluding them solely because their spreading is less aesthetically pleasing.

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A learner claims that 20 normal metaphases exclude a 10% abnormal population. Calculate approximately a 12.2% chance of missing it under ideal sampling, then consider whether culture bias makes the real limitation even greater.

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A worksheet lists 20 counted cells but only a subset with complete band-by-band analysis. Describe those activities accurately rather than stating that all 20 cells were fully assessed for subtle structural abnormalities.

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Two objectives have the same magnification but different numerical apertures. For resolving closely spaced band detail under suitable illumination, favor the higher numerical aperture objective and ensure its required immersion medium and optical conditions are correctly used.

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A fluorophore is bright on a verified compatible microscope but nearly invisible after a filter cube change. Check excitation, dichroic and emission compatibility before repeating specimen processing or calling the target signal absent.

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A marrow shows a small derivative chromosome 22 and a corresponding chromosome 9 rearrangement. Recognize the classic Philadelphia pattern and correlate it with targeted BCR::ABL1 testing rather than inferring molecular identity from size alone.

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A rearranged chromosome retains the chromosome 4 centromere but carries distal chromosome 8 material. Identify it as a derivative chromosome 4, then evaluate which chromosome 4 material is missing and which chromosome 8 material is additional.

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One homolog has a larger heterochromatic region but otherwise matching euchromatic bands. Evaluate whether it fits a recognized variant before ordering broad conclusions about pathogenicity, while investigating any atypical band change beyond the expected polymorphic region.

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Automated chromosome separation appears to remove a small segment from an overlapping pair. Inspect the original image and revise segmentation using the actual boundaries rather than accepting the edited karyogram as evidence of a deletion.

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Worked teaching report: Bone marrow investigated for a suspected myeloid neoplasm. G-banded analysis: 20 metaphases counted and fully analyzed at approximately 400-band resolution; three representative karyograms prepared. Result: 47,XY,+8[12]/46,XY[8]. Annotation: 47 is the total chromosome number in the abnormal cells; XY describes the sex chromosome complement; +8 identifies an additional chromosome 8; [12] and [8] give the numbers of metaphases with each finding. Interphase FISH on uncultured marrow: nuc ish (D8Z2)×3[120/200]. Annotation: nuc ish identifies interphase nuclear hybridization; D8Z2 identifies the chromosome 8 centromeric target; ×3 means three signals; [120/200] means that pattern occurred in 120 of 200 scored nuclei. The remaining 80 nuclei had two signals. With acceptable controls and a result exceeding the validated cutoff, interpretation is: a trisomy 8 clone is detected, with concordant targeted FISH findings. Trisomy 8 alone does not establish a specific myeloid diagnosis; correlate with morphology and molecular findings.

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A marrow accession is divided among culture, direct FISH and DNA extraction. Ensure each derived container and record retains a verified link to the accession, and document transfers before the branches enter separate work areas.

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A colcemid container has acceptable dating but an unexpected change in appearance and a questionable storage history. Remove it from use pending assessment and review affected preparations instead of relying solely on the printed expiration date.

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An incubator display remains normal, but an independent check shows a sustained temperature deviation and several cultures grow poorly. Investigate the excursion, assess affected specimens and verify restored performance before returning the unit to routine use.

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A worker plans to prepare volatile methanol–acetic acid fixative inside a recirculating cabinet used for cell culture. Select appropriate chemical ventilation rather than assuming that protection against biological aerosols also controls accumulated solvent vapors.

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Before introducing a new fixative component, review its safety data sheet and determine whether the Chemical Hygiene Plan addresses its ventilation, protective equipment, storage and spill response needs. Update procedures and training where gaps exist.

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After changing a banding procedure, the laboratory records fewer poorly banded slides but more repeat preparations. Review both indicators to determine whether the change improved overall performance rather than celebrating one isolated metric.

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A technologist recognizes a patient's name and wants to examine the chromosome report despite having no role in that case. Access should be based on an authorized work need rather than personal familiarity or interest.