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The HMDS Department

Introduction
Description of the Service
Quality Assurance and Audit
Funding and Costs
Sending Specimens to HMDS
Postal Address
Request Form


Introduction

The Haematological Malignancy Diagnostic Service was formed in 1993 by the amalgamation of the Leukaemia Diagnostic Unit based at Cookridge Hospital, and the Yorkshire Regional Cancer Organisation Histopathology laboratory based at Leeds General Infirmary.

Aims of the Service

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Description of the Service

The diagnosis of the various types of haematological malignancy requires expertise in the microscopic examination of cells and tissues, the application of cell marker studies and the use of molecular techniques. The main workload is concerned with the investigation of peripheral blood, bone marrow and lymph node specimens. The methods used can be applied to any sample of tissue or cell suspension where there is clinical suspicion of haematological malignancy.

Histology

The laboratory will investigate any type of biopsy or surgical specimen in which haematological malignancy is suspected. Most of the histology workload consists of lymphoid tissue, bone marrow and skin biopsy specimens. All bone marrow and skin biopsy specimens are processed in methyl methacrylate resin. This resin has the unique property of combining excellent morphological detail with the ability to carry out a wide range of immunohistological marker studies making it the method of choice for the examination of bone marrow biopsy specimens.

The routine use of antigen retrieval techniques has greatly expanded the range of immunohistological markers which can be performed on fixed tissue embedded in paraffin or resin. HMDS now has in excess of 60 markers which can be used on fixed tissue. A further selection of antibodies are available for use on cryostat section when fresh tissue is available. Unless there are specific problems most immunohistological studies are completed in 24 hours after a preliminary H & E section has been examined.

Immunohistological studies are carried out using panels designed for the investigation of specific disorders (eg suspected Hodgkin's disease, cutaneous T-cell lymphoma, etc), and provide an essential cross check on the validity of a morphological diagnosis. The use of panels of this type also allows effective auditing of the performance of individual antibodies. HMDS is currently carrying out around 15,000 immunohistological investigations per year making it one of the busiest laboratories in the United Kingdom for this type of work.

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Flow Cytometry

The investigation of cell suspensions of blood, bone marrow, cerebrospinal fluid, stem cell harvests and disaggregated tissue by flow cytometry is the largest single activity in the laboratory. The laboratory uses an upgraded Becton Dickinson FACScan and a FACSort. Approximately one hundred antibodies are available for flow cytometric investigations. These include more than one antibody from many of the common CD categories which can be used to follow up unexpected results.

All flow cytometry studies are carried out using combinations of two, three or four antibodies labelled with different fluorescent dyes. This is essential to ensure that results can be accurately reported for the abnormal cell population in a sample that consists of a complex mixture of cell types. In this application the use of a single antibody technique is now obsolete. All the antibodies are used in preset panels chosen to give the maximum diagnostic information.

The ability to sort and collect small populations of cells greatly enhances the ability to detect minimal disease. The technology of flow cytometry is now very reliable and sophisticated. However, the investigation of haematological malignancy requires considerable operator expertise. An annual workload of about 65,000 tests ensures that staff maintain a high level of experience. Flow cytometry is a rapid technique and in urgent cases results can be obtained in 1-2 hours.

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Molecular Biology Techniques

Molecular biological techniques are now an integral part of the laboratory diagnosis of haematological malignancy. These techniques allow the investigation of T and B-cell monoclonality and the detection of an increasing number of chromosomal abnormalities. Most of the work is carried out using polymerase chain reaction (PCR) based methods. These are rapid and allow the use of poor quality DNA such as may be obtained from fixed tissue or blood smears. Many of the PCR techniques use fluorescent primers in the Applied Biosciences Genescan system which is a highly sensitive detection system. Quantitative PCR methods which allow an assessment of the extent of residual disease in CML are now in routine use.

PCR methods are now complemented by in situ hybridisation techniques which are of particular value in detecting abnormal chromosome numbers such as monosomy 7 or trisomy 12 and for the detection of some chromosomal translocations in both fresh and fixed cells.

Tests for a number of common clinically important chromosomal abnormalities are now routinely performed in cases of acute leukaemia and lymphoma.

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Monoclonal Antibody Production

If HMDS purchased all the monoclonal antibodies required from commercial sources the annual cost would exceed £100 000. Fortunately, about 60% of the antibody requirement is produced within the laboratory. A wide range of monoclonal antibodies are produced from cultured hybridoma cells, purified and conjugated with the fluorochromes FITC, PE, Cy5 and APC. These antibodies have also been tested in other laboratories and have been found to be of high quality. The cost of production, including all overheads, is 80-90% less than the price charged by commercial suppliers.

The rapid development of areas such as stem cell manipulation and drug sensitivity testing means that cell culture based activities may become an increasingly important part of the work of the laboratory in the years ahead. A number of aspects of this work are under development.

Research and Development

Research is an essential component of the service and ultimately benefits the users of the service. Major research activities are funded by grants from the Leukaemia Research Fund, Yorkshire Cancer Research Campaign and Friends of the Leukaemia Unit at Leeds General Infirmary.

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Quality Assurance and Audit

Audit within the laboratory has a high priority and has a number of main components:

Within the General Infirmary there is a very close relationship with the Clinical Haematology Unit which is facilitated by several members of staff who work in both the laboratory and in the clinical area.

HMDS has Clinical Pathology Accreditation (CPA) and participates in a range of technical external quality assurance programmes:

Funding and Costs

The main workload of the laboratory originates from hospitals in North, West and South Yorkshire; this represents a population of 3.5 million. The work is funded through block contracts with NHS purchasing authorities allowing open access to the service. Work from other NHS units or private hospitals is charged on a cost per item basis. Current charges are available on request.

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Sending Specimens to HMDS

Tissue Specimens (excluding peripheral blood and bone marrow)

Ideally lymph node biopsies and other tissue specimens should be sent to the laboratory as unfixed tissue. This allows marker studies to be carried out by flow cytometry and improves quality of material prepared for FISH or PCR based investigations.

The portion of the specimen to be used for histology and immunocytochemistry can be processed under standardised conditions to ensure optimal results. Specimens should be sent in tissue culture medium or sterile saline by the fastest available method of transport. Where no unfixed tissue is available tissue in 10% formalin or paraffin blocks may be sent.

Bone Marrow

Where possible the specimen should include freshly prepared unstained smears, a sample of bone marrow in EDTA and a trephine biopsy in 10% formalin. The specimen should be received within 24 hours. However, if RT-PCR may be required, as part of the diagnosis of acute leukaemia, the specimen should be sent by the fastest available route.

Blood

10mls of blood should be sent in EDTA to arrive within 24 hours.

Effusions and CSF

These do not generally require anti-coagulants and should be sent in a sterile container.

Postal Address

HMDS
Level 3, Bexley Wing
St. James's Institute of Oncology
Beckett Street
Leeds LS9 7TF

Fax: 0113 206 7883

HMDS request form:

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Document last updated: Wednesday, 30 September 2009

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