Cytogenetics

Postnatal diagnosis (blood culture), prenatal diagnosis; solid tissue, FISH, and cell line karyology.

Cytogenetic analysis is performed according to the Professional Guidelines for the Association of Clinical Genetic Science, and the recommendations provided are dependent on the clinical indications given for each case.

Clinical details

Clinical details are very important when providing genetic analysis. The more clinical information that is available (e.g. details of ultrasound information, phenotypic features or family history), the better the service we can provide. Failure to provide this information for cytogenetic studies may result in an inaccurate analysis.

Clinical details inform the investigation at all stages:

• Prior to analysis, clinical details may indicate, for example, that procedures such as chromosome breakage or leukaemic studies are required, which must be referred to  the oncogenomic department or specialist centre.
• During analysis, they may indicate that extra cells should be screened to investigate the possibility of mosaicism (in a diagnosis of suspected Turner syndrome, for example), or that particular chromosomes must be targeted for high-resolution study (chromosome 4 in suspected Wolf-Hirschhorn syndrome, for example).
• When the analysis has been completed, they may help to provide accurate interpretation of the findings and, in some instances, prompt further investigations – FISH or molecular genetic studies, for example.

When clinical details are not available, a routine analysis will be performed and a conditional report issued.

Cytogenetic sample stability

Cytogenetic studies require living cells. Please ensure that samples reach the laboratory as soon as possible. If a delay before dispatch is unavoidable, samples may be stored in a refrigerator (4°C) but they must not be frozen.

Samples sent more than 48 hours after sampling, or kept at temperatures below 4 ̊C and greater than 38 ̊C, may have inhibited growth.

Information concerning packaging, transportation, and labelling of samples is provided on the reverse of our TDL Genetics Request Form.

Postnatal diagnosis (blood culture)

Reasons for analysis: Chromosome studies are requested where problems that may have a cytogenetic basis are suspected, e.g. babies with birth defects; children with developmental delay and physical handicaps, or adults with fertility problems. Additionally, prospective gamete donors are screened to detect carriers of balanced chromosome rearrangements.

Sample requirements: Lithium heparin whole blood specimens are required – gently mixed to prevent clotting and must not be frozen. See sample stability section for cytogenetic samples. Sample volumes may be reduced for children (2–4ml) and neonates (1-2ml).

Turnaround time: The usual turnaround time is 2–3 weeks; however, the laboratory will endeavour to respond to urgent requests. Where a major trisomy is suspected, a rapid PCR screen may be performed to provide an urgent provisional result.

Notes

• Rarely, blood samples fail to culture (<1%).
• The culture may yield chromosomes of insufficient quality. This will be indicated on the report and a repeat study suggested.
• The laboratory should be informed if the patient has recently received a blood transfusion.
• The laboratory should be informed if the patient has EVER had a bone marrow transplant.
• The patient’s biological sex should be included on the request form.

Prenatal diagnosis

Reasons for analysis: Chromosome studies are requested where pregnancies are identified as being at risk of a cytogenetic abnormality e.g. advanced maternal age; positive maternal serum screening; fetal abnormalities found on ultrasound; or where a parent is a known carrier of a chromosome anomaly, or where a high risk trisomy has been found by NIPT. 

Sample requirements:

• Amniotic fluid – 10ml+ in a plain sterile, leak-proof container. Suitable containers can be provided by the laboratory. The specimen must not be frozen. See sample stability section for cytogenetic samples.
• Chorionic villus – 5mg+ in sterile transport medium. Suitable containers containing medium can be provided by the laboratory. The specimen must not be frozen. See sample stability section for cytogenetic samples.
• Fetal blood–1–2ml LITHIUM HEPARIN whole blood, gently mixed to prevent clotting. The specimen must not be frozen. See Cytogenetic sample stability above.

Turnaround time: This is dependent on the rate of cell growth, however, the usual turnaround time is approximately 2 weeks. A number of circumstances now occur more frequently, as invasive prenatal diagnosis becomes less common, which may result in delayed reporting time. These include:

• A delay in transportation in order to collect a batch of samples to reduce courier costs. Even when couriered promptly, sample growth may be slower than that seen in samples sent immediately.
• Sampling at early or late gestations, for example to confirm non-invasive tests or follow up anomaly scans.
• A tendency to take smaller quantities of sample or to take insufficient sample for multiple techniques.
• The request for karyotyping as an add-on after an initial PCR test.

Fetal blood results will usually be reported by 10 calendar days. For all other prenatal tests, please contact the laboratory prior to taking samples.

Notes

• Maternal contamination, and mosaicism may complicate the analysis and may lead to the suggestion that a second invasive test is performed.
• Rarely, cultures fail to grow (overall <1%).
• Very small chromosome abnormalities may not be detected (this is why the phrase ‘No trisomies or major chromosome abnormalities detected…’ is used in our reports).
• For TTTs or heavily blood stained amniocentesis samples, please provide a maternal EDTA blood sample for comparison studies.

Solid tissue

Reasons for analysis: Fibroblast cultures may be used in addition to blood cultures, for example where tissue specific mosaicism is suspected, or where blood samples cannot be obtained. POC samples may be requested for early spontaneous miscarriages, stillbirths, or to confirm a prenatal diagnosis.

Sample requirements: All specimens should be placed in a sterile container, preferably containing transport medium. This can be supplied by the laboratory. Sterile normal saline can be used if transport medium is not available. Samples must not be placed in formaldehyde or other preservative and must not be frozen. See sample stability section for cytogenetic samples.

Turnaround time: This is dependent on the rate of cell growth, however, the usual turnaround time is approximately 4 weeks.

Notes

• Material from miscarriages has a relatively high culture failure rate (around 20%). Where failure occurs, alternative molecular methods may be attempted, usually a KaryoLite Bacs-on-Beads assay that can detect whole monosomy or trisomy of any chromosome, if possible.
• If no villus or fetal parts are identified in supposedly POC material and a normal female chromosome result is found, this may indicate that maternal tissue has been cultured (this will be noted on our report).
• If a request is made for remaining pregnancy loss tissue to be returned to the patient or hospital for burial or cremation, we will return the sample as soon as possible once adequate tissues have been used for testing. Please ensure that this is communicated to the lab using a hospital consent form, noted on the referral form or by email. Patients can arrange to collect remaining tissues from TDL Patient Reception.
• For samples without specific consent, the lab will send all remaining tissue for sensitive incineration. Please note that there is no distinction made between fetal and other pregnancy tissues for this process, and there will be no ashes afterwards. The lab keeps detailed records of all pregnancy tissue sent for incineration, and a Certificate of Destruction is available if required.

Fluorescence in situ hybridisation (FISH)

Where FISH studies for specific microdeletion syndromes are required, this must be indicated on the request form.

Note: FISH studies for a rapid pre or postnatal aneuploidy screen have now been superseded in our laboratory by multiplex-PCR technology. Subtelomeric screens are now performed by Array CGH as part of developmental delay investigations. Common microdeletion syndrome testing is now performed by BOBs analysis.