Haematology in under-resourced laboratories

Chapter 26 Haematology in under-resourced laboratories



Imelda Bates, Jane Carter



Chapter contents


Introduction: types of laboratories 603

Organization of clinical laboratory services 604
Level A: Sub-district facilities including health centres 604

Level B: District hospitals 604

Level C: Central and teaching hospitals 605

Availability of tests at each level 605
Level A 605

Level B 605

Level C 605

Microscopes 606
Care of the microscope 606

‘Essential’ haematology tests 606
Cost per test 606

Diagnostic reliability 606

Clinical usefulness 607

Maintaining quality and reliability of tests 607
Quality control of a test method (technical quality) 607

Internal quality control 607

External quality assessment 607

Basic haematology tests 607
Haemoglobinometry 607

Direct reading haemoglobinometers 608

Haemoglobin colour scale 608

Packed cell volume 609

Manual cell counts using counting chambers 609

Total white blood cell count 610

Platelet count 610

Peripheral blood morphology 612

Modified (one-tube) osmotic fragility test 612

Haemoglobin E screening test 613

Tests for paroxysmal nocturnal haemoglobinaemia 613

Reagents for prothrombin time (PT) and activated partial thromboplastin time (APTT) 613

Laboratory support for management of HIV/AIDS: CD4-positive T-cell counts 614

Laboratory management 614
Interlaboratory communication 614

Specimen transport 614

Staff training 614

Clinical staff interaction 615

Health and safety 615


Introduction: types of laboratories


In most countries, there are likely to be some laboratories with limited resources, but in under-resourced countries, the majority of laboratories face chronic shortages of trained staff, low morale, inadequate equipment and erratic supplies of reagents. These have a major impact on the range and quality of services offered. Many laboratories lack the highly sophisticated equipment found in more wealthy settings and still operate using manual techniques. Smaller laboratories tend to be multifunctional, performing a range of haematology, parasitology, clinical chemistry and microbiology tests. A blood transfusion service is usually available at larger institutions and, unless there is access to a national blood service, laboratory staff are responsible for donor selection, blood collection and issuing of blood. If there is no separate system of public health laboratories, clinical laboratories may also be required to provide reliable health surveillance data for epidemiological and public health monitoring and to investigate disease outbreaks and refer samples for confirmation.


In under-resourced countries, these difficulties are compounded by the high burden of infectious diseases such as HIV/AIDS, tuberculosis and malaria. There are also additional pressures from external agencies for diagnostic and monitoring services which may not accurately reflect local public health priorities. Parasitological diagnosis of malaria is now recommended in all age groups to prevent the over-diagnosis and misuse of new anti-malarial combination drugs. Although it is currently the subject of much debate, WHO guidelines state that the only exception to the requirement for parasitological diagnosis is children aged under 5 years living in stable high-transmission settings where the likelihood of malaria as a cause for fever is high.1 Occasionally, in tertiary centres, the diagnosis of tuberculosis requires aspiration and culture of the bone marrow and trephine biopsy examination, especially in patients who are HIV positive, in whom sputum tests for acid-fast organisms are frequently negative.2 The decision to initiate antiretroviral therapy and the monitoring of therapy require regular haemoglobin concentration (Hb) estimations, CD4-positive lymphocyte counts (or percentages for paediatric care) and, ideally, plasma viral load determinations, although the maintenance of equipment for some of these tests is challenging for low-income countries.3,4


The purpose of this chapter is to provide guidance for an effective haematology service at the different levels of the healthcare system in resource-limited countries. In planning such services, it is necessary to determine what tests are needed at each level and to identify the relevant referral networks for clinical haematology. For example, successful management of haematological malignancies in countries with limited resources might involve partnerships between local institutions and those in more wealthy countries to adapt treatment protocols, to provide training or diagnostic services or to improve local supportive care facilities.5,6



Organization of clinical laboratory services


In under-resourced countries clinical laboratory services may be considered at three levels according to their size, staffing and services offered. These are levels: (A) subdistrict facilities including health centres; (B) district and referral hospitals; and (C) central reference and teaching hospitals (Fig. 26.1).


image

Figure 26.1 Network of laboratories.



Level A: Sub-District Facilities Including Health Centres


The level A laboratory provides basic diagnostic services to support patient management on-site and determine when a patient requires referral. In some countries, there may be no formal laboratory and some diagnostic tests are carried out at the point-of-care by nurses, assistants or orderlies with no technical qualifications. Haematology equipment at this level includes a method for estimating Hb and a microscope for examining slides for tuberculosis and malaria. However, maintenance of equipment in rural areas is often difficult and technical work may not be properly supervised, which can significantly compromise the quality of results.



Level B: District Hospitals


District hospital laboratories are usually multipurpose, performing microbiological and biochemical as well as haematological tests. Laboratory staff may comprise one or two qualified biomedical scientists supported by assistants who often have little or no formal training but have learnt various techniques at the bench. Equipment available for haematology may include a microscope and centrifuge and possibly a basic colorimeter for measurement of Hb. In some district hospitals, donor funds have provided more sophisticated equipment such as automated haematology and clinical chemistry analysers, predominantly to support HIV services. In the absence of a national blood service, district hospitals are responsible for blood transfusion services so the laboratory will be expected to perform blood grouping and crossmatching as well as screening for at least HIV, hepatitis B virus and possibly hepatitis C virus.



Level C: Central and Teaching Hospitals


At this level, the laboratory staff have usually received multidisciplinary training and some of the senior staff have received postgraduate training in a specialist laboratory discipline. Each laboratory usually has a specialist technical head who works closely with the clinical pathologist. Equipment available includes centrifuges, colorimeters, microscopes, haemoglobin electrophoresis equipment and possibly blood bank centrifuges for the separation of blood components. Automated haematology analysers are usually found in such laboratories. In many cases, these have been supplied by donor agencies, but long-term funding to support maintenance of these systems is often lacking and, consequently, they may be unreliable or not used at all owing to a shortage of appropriate reagents and inadequate maintenance.



Availability of tests at each level


In under-resourced countries, the haematology tests available at the different levels of healthcare vary widely and depend on local clinical needs as well as availability of equipment and qualified technical laboratory personnel. The following is a general description of the tests that are likely to be required at each level.



Level A



• Haemoglobin estimation by a manual method (see p. 608)

• Malaria testing on peripheral blood thick and thin films (see p. 109) or rapid diagnostic test for Plasmodium falciparum and other species (see p. 110)

• Testing for HIV infection.


Level B



• Haemoglobin measurement (see p. 24)

• Peripheral blood morphology, especially to identify the cause of anaemia (Chapter 5)*

• Total white blood cell counts (see p. 610)

• Differential white cell count (see p. 31)

• Platelet estimates (usually from blood film)

• CD4 lymphocyte count (see p. 614)

• Malaria testing by thick and thin peripheral blood films (see p. 109) or rapid diagnostic test for Plasmodium falciparum and other species (see p. 110)

• Screening test for sickle haemoglobin in areas where this is relevant (see p. 314)

• Blood grouping and compatibility testing (Chapter 21)

• Some larger laboratories may have automated (Hb concentration), mean cell volume (MCV), mean cell haemoglobin (MCH), mean cell haemoglobin concentration (MCHC), white blood cell total and differential counts, platelet counts (see p. 36).


Level C


In addition to tests carried out at level B, the haematology services offered by level C may include the following:


• Automated Hb, MCV, MCH, MCHC, white blood cell total and differential counts, platelet counts (see p. 36)

• Haemoglobin electrophoresis or high-performance liquid chromatography (Chapter 12)

• Haemoglobin A2 and haemoglobin F measurements (see pp. 322 and 325)

• Glucose-6-phosphate dehydrogenase screen (by fluorescent spot or methaemoglobin reduction method) (see p. 255)

• Flow cytometry immunophenotyping (Chapter 14)

• Polymerase chain reaction (PCR) or other method for diagnosis of mutations associated with haematological malignancies (Chapter 21)

• HIV plasma viral load estimations

• Staining of bone marrow smears for morphological assessment (see p. 59) and estimation of iron status (see p. 333)

• Bone marrow trephine biopsy examination (see p. 123)

• Identification of blood group antibodies (Chapter 19)

• Basic clotting screen (prothrombin time, thrombin time and activated partial thromboplastin time) (see p. 409)

• Oral anticoagulant control (see p. 467)

• Separation of whole blood into packed cells, plasma and, occasionally, platelets.


Microscopes


The microscope is the most important piece of laboratory equipment in under-resourced countries and is essential for the diagnosis of anaemia, malaria and other blood parasitic infections and for performing total and differential white blood cell counts. Reliable assessment of morphological features requires a microscope that is clean and correctly set up with aligned lenses and an electric light source (either inbuilt or reflected light) to ensure clear images, especially at high magnification. Failure to maintain microscopes to a high standard by routine user maintenance or, ideally, with regular professional servicing, can lead to inaccurate diagnoses and inefficient use of technician time.8,9 Routine maintenance of the microscope is described on p. 52.



Care of the Microscope


In hot, humid climates, if no precautions are taken, fungus may develop on the microscope – particularly on the surface of the lenses, in the grooves of the screws and under the paint – and the instrument will soon be useless. This can be prevented by placing the microscope every evening in an airtight dust cover together with silica gel. Dry the silica as necessary and reuse it. An alternative method is to place the microscope in a warm cupboard with a tight-fitting door, heated by a 40-watt light bulb. Check that the temperature inside the cupboard is at least 5°C warmer than that of the laboratory, but take care that it does not overheat.


In hot, dry climates, the main problem is dust. Fine particles work their way into the threads of the screws and under the lenses. This can be avoided as follows:


1. Always keep the microscope under a dustproof plastic cover when not in use.

2. At the end of the day’s work, clean the microscope thoroughly by blowing air onto the lenses and moving parts using a rubber bulb.

3. Finish cleaning the lenses with a lens brush or fine paintbrush. If dust particles remain on the surface of the objectives, remove with clean lens paper.


‘Essential’ haematology tests


Despite the relatively high cost of running a laboratory service and the low per capita healthcare budget in under-resourced countries, there are few data available on which to base rational decisions about ‘essential’ laboratory tests.10,11 In many of these countries, decisions are made at central level by healthcare planners without adequate consultation with laboratory managers and experts. The selection of ‘essential’ tests at each level should be based on the clinical and public health needs of the local community and the availability of qualified clinical and laboratory staff, as well as on the availability of funds. Essential test packages are usually defined as part of national policy and standards, taking into consideration medium- and long-term trends and the requirements of disease control programmes.


To ensure cost-effectiveness of the laboratory service, tests with no proven value should be eliminated and new tests for which there is independent evidence of clinical usefulness should be introduced,12 as described in Chapter 24, p. 567. Tests that provide objective qualitative or quantitative information are preferred. Although it is not possible to draw up a list of essential tests that will be applicable to all countries, or even to different regions within a country, the following aspects should be considered.



Cost per Test


Often the cost of a test is calculated from the price of reagents divided by the number of tests performed. However, this oversimplifies the situation and is not accurate enough to form the basis for national policy decisions and budget allocation.11 The factors that need to be taken into account when calculating the total annual costs for a laboratory are given in Chapter 24.



Diagnostic Reliability


It is important to know the sensitivity and specificity of a test and its predictive values (as calculated on p. 567) when selecting a laboratory test for clinical use. This information may be provided by manufacturers but it may not be locally applicable and, in many under-resourced countries, local test evaluation data are not available. In some countries, the ‘gold standard’ diagnostic services needed to determine ‘true positive’ and ‘true negative’ data in the local context are also lacking.


The quality of all tests carried out by a laboratory should be regularly monitored; systems for doing this are well-established (see Chapter 24) but are not easily implemented in under-resourced settings. The quality of a test influences its usefulness as well as its utilization by clinicians and community members. For example, if the result of a test in routine practice is correct only 80% of the time, then one in five tests will be wasted, reducing the effectiveness of the test by 20%. Furthermore, the inaccurate test may result in a patient receiving inappropriate treatment. Clinicians and the general public are increasingly aware of the need for accurate diagnosis. Clinicians may not order tests or use the results in patient management decisions if they do not trust the quality of the laboratory results13 and poor quality healthcare may deter patients from using health facilities.



Clinical Usefulness


An assessment of the clinical usefulness of a test should be carried out by an independent clinician who is familiar with local diseases and the diagnostic support services that are available. This assessor needs to compare actual clinical practice with locally agreed ‘best practice’ or, if available, local guidelines.12 From observation of a range of clinical interactions, the percentage of times that ideal practice is followed can be calculated. For example, transfusion guidelines may recommend that transfusions are given routinely to children with an Hb of <5 g/dl. The assessor can record how many children with Hb below this level failed to receive a transfusion and how many transfusions were given without waiting for the Hb result or at an inappropriate Hb level. For each test, the assessor needs to judge whether the test has been appropriately requested and is used to influence patient management or public health decisions. The percentage of tests that are not used to guide clinical decisions will provide a figure for ‘clinical wastage’ of the test that can be entered into the formula (Chapter 24).



Maintaining quality and reliability of tests


Paradoxically, it is in under-resourced laboratories, where equipment and supplies are limited and training and supervision may be minimal, that the level of skills and motivation required to maintain a good-quality service need to be highest. Even the most basic of laboratories should ensure that procedures are in place to monitor quality (see Chapter 25). In addition to monitoring the technical quality of each test, the quality of the whole service must be ensured both within the laboratory and between laboratories. Standard operating procedures (see p. 575) should be drafted for every method performed in the laboratory; these can be adapted from existing standard operating procedure ‘models’. In addition to providing standardized techniques, standard operating procedures are excellent teaching resources and adherence to these procedures will minimize errors. Standard operating procedures need to be regularly reviewed and updated to keep pace with technical developments and changes in local circumstances (e.g. non-availability of reagents, technical limitations).



Quality Control of a Test Method (Technical Quality)


Methods for the control of various haematology tests are described in Chapter 25 but some may need to be adapted to specific local circumstances in resource-poor countries. For example, if commercial controls are not affordable, each batch of sickle cell screening tests should include known positive and negative samples from a previous batch of tests; for monitoring constancy of Hb estimations, a high and a low value sample can be re-tested several times during the day.



Internal Quality Control


Internal quality control is a system within an individual laboratory for ensuring that the technical elements of the test are of acceptable quality. Monitoring of quality by processing control samples and plotting a control chart (see p. 593) will highlight major problems with the system. For example, an inaccurate differential white cell count may be due to problems with sample collection and handling, slide preparation, fixing and staining, morphological interpretation and microscope quality as well as inadequate microscopy technique. Measures such as the introduction of standard operating procedures, in-service training and equipment maintenance schedules can help to improve performance and reduce inaccuracies.



External Quality Assessment

Related posts:

Reference ranges and normal values Quality assurance Laboratory control of anticoagulant, thrombolytic and antiplatelet therapy Molecular and cytogenetic analysis Bone marrow biopsy Erythrocyte and leucocyte cytochemistry

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Jun 12, 2016 | Posted by in HEMATOLOGY | Comments Off on Haematology in under-resourced laboratories

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