Within NHS Lothian, diagnostic BCR-ABL1 reverse transcription PCR (RT-PCR), and BCR-ABL1 real time PCR (QPCR) for minimal/ measurable residual disease quantification analysis can be requested electronically via the TRAK system.
Requests for testing on formalin-fixed paraffin embedded (FFPE) tissue should be sent to the Molecular Pathology Service for extraction, along with a completed HMDS test request form.
For all other requests a specimen should be sent to the Molecular Haemato-Oncology laboratory along with a completed request form (above) following local protocols for the transport of biological samples. Requests for myeloid Next Generation Sequencing testing should use the NGS request form.
Please ensure the request form is completed fully; in particular three points of patient identification are required including the CHI number when possible.
All specimen containers should be fully labelled and securely packaged. It is recommended that centres referring cases send an email notification to HMDS.Lothian@nhslothian.scot.nhs.uk.
The Molecular Haemato-Oncology Laboratory is funded by NSD Scotland; all tests are provided free of charge for requests received from within NHS Scotland.
Specimen Requirements
Tests are routinely performed on DNA and RNA extracted from peripheral blood or bone marrow specimens. It is essential that all specimens are collected in are routinely performed on DNA and RNA extracted from peripheral blood or bone marrow specimens. It is essential that all specimens are collected in EDTA tubes to enable molecular testing.
Many of the assays are performed using RNA as a template, due to the labile nature of RNA, specimens should ideally arrive in the laboratory within 24 hours of collection (maximum of 48 hours). Specimens should therfore not be sent on Fridays where possible; if this is unavoidable please telephone the laboratory and discuss with a member of our team.
Please see individual test details information in the test directory for details on required sample volumes.
Where testing is to be performed on DNA or RNA extracted from FFPE tissue samples, tissue blocks containing sufficient neoplastic cells for the individual tests should be provided (normally 10% neoplastic cell content is required). The corresponding H&E slide and a copy of the pathology report should also be provided.
Where testing is to be performed on DNA or RNA already extracted from FFPE tissue, the sample should be sent along with the corresponding pathology report and an estimated tumour percentage .
Factors known to affect quality of molecular testing:
- Transport time: RNA is labile and therefore RNA extractions must be performed as quickly as possible to prevent degradation. Samples should ideally arrive in the laboratory within 24 hours of collection. Where possible, samples should not be sent so that they arrive late on Fridays.
- Transport temperature: Due to the labile nature of RNA, specimens should be sent at room temperature to prevent degradation and a decrease in sample quality.
- Sample volume: For RNA based assays, a large sample volume is often needed to obtain the required RNA content for downstream molecular testing. For further details on required sample volumes please see individual tests in the test directory.
- Specimen type: Whenever possible please provide the specimen type advised in the test directory to ensure testing can be performed.
- Collection tube: All blood and bone marrow specimens must be collected in EDTA tubes. Specimens not collected in EDTA tubes will not be suitable for molecular analysis. Clotted samples are not suitable for molecular analysis.
- Cell yield: Samples with a low cell count may yield inadequate quantity of nucleic acid for analysis.
- FFPE tissue quality: Tissue fixation quality, including ischemic time, duration of fixation, sample size (i.e. penetration of formalin) and processing protocols can affect the quality of DNA isolated from FFPE tissues, and therefore the results of molecular analysis.
- Acid decalcification of tissues (e.g. using formic acid or hydrochloric acid-based decalcifying agents) is known to degrade DNA in FFPE samples making subsequent molecular analysis impossible
- The use of the plasma-thrombin technique for the preparation of cell blocks from cytology specimens can introduce DNA contamination into a sample and is strongly discouraged unless the plasma has been DNase treated
- Non-neoplastic cells within a sample may dilute the analyte of interest beyond the level of detection of some molecular diagnostic assays.
If analysis is required of a specimen type not described above, please contact a member of the Molecular Haemato-Oncology team prior to requesting.
Mandatory Data Set
Referring clinicians are requested to comply with NHS Lothian policy on mandatory data set requirements for patient identification on laboratory requests. The following information must be legible on the request form:
- Patient identifier number (CHI or NHS number)
- Surname
- Forename
- Date of Birth
- Gender
- Location/Requesting Clinician
If the CHI/NHS number is not available, the data set must include first line of the patient’s address and postcode.
Tissue blocks sent from external laboratories must be labeled with a unique identifier. Accompanying slides must be labelled with the unique identifier and patient surname.
Failure to comply with this policy will result in the sample being rejected. Samples without minimum data, or for which data on the sample and accompanying documents do not match, will not be accepted and will be returned to the sender.
Consent, Storage and Authorisation
In accordance with the requirements of the Human Tissue (Scotland) Act 2006, it is the responsibility of the referring clinician to ensure that appropriate informed consent has been obtained before any testing is undertaken. The laboratory must be informed of any restrictions to this consent (e.g. storage of samples).
Unless otherwise informed, the laboratory assumes that all appropriate consent has been obtained from the patient for the tests requested and for storage of the derived DNA or RNA (cDNA) for future use both in assisting further testing (if required) and in the development of future diagnostic tests.
If in doubt, contact a member of the Molecular Haemato-Oncology team to discuss.
Specimen Transportation
It is the responsibility of those dispatching specimens to the laboratory to ensure that these samples are sent in accordance with any national guidelines and/or local policies for the packaging, labelling and transport of biological material.
The Health and Safety Executive guidance, “Biological Agents: Managing the risks in laboratories and healthcare premises”, recommends materials such as blood, tissue, excreta, secreta etc collected from humans be considered, as a minimum, Category B infectious substances.
The following packaging, labelling and transport requirements are applicable for samples from otherwise healthy individuals, or where there is no reason to suspect that they are suffering from a severe infectious disease, and are derived from Packing Instruction P650 of the European Agreement concerning the International Carriage of Dangerous Goods by Road (ADR). Such specimens are not subject to ADR provided the specimen is carried in a packaging which will prevent any leakage and which is marked with the words "EXEMPT HUMAN SPECIMEN".
The packaging is deemed to comply with the above requirements if it meets the following conditions:
- The packaging should consist of three components: leak-proof primary receptacle(s), a leak-proof secondary packaging, and an outer packaging of adequate strength for its capacity, mass and intended use, and with at least one surface having minimum dimensions of 100 mm × 100 mm.
- For liquids, absorbent material in sufficient quantity to absorb the entire contents should be placed between the primary receptacle(s) and the secondary packaging so that, during carriage, any release or leak of a liquid substance will not reach the outer packaging and will not compromise the integrity of the cushioning material.
- When multiple fragile primary receptacles are placed in a single secondary packaging, they must be either individually wrapped or separated to prevent contact between them.
Packages should be clearly labelled with the delivery address and sender details. Labels must be durable and legible.