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Immunophenotyping by Flow Cytometry

Acknowledgement The authors would like to acknowledge Kathryn Moss for her contribution to the HIV monitoring section. Principles of flow cytometric immunophenotyping The primary methods for immunophenotyping in the haematological setting are immunocytochemistry (described in previous editions of this book) and flow cytometry. In this chapter we shall focus on flow cytometry and its application to the diagnosis and treatment response of haematological neoplasms. Flow cytometry…

Erythrocyte and Leucocyte Cytochemistry

Acknowledgement The major contribution of the late Dr David Swirsky to this chapter in previous editions of the book is gratefully acknowledged. Erythrocyte cytochemistry Siderocytes and sideroblasts Siderocytes are red cells containing granules of non-haem iron. They were originally described by Grüneberg in small numbers in the blood of normal rat, mouse and human embryos and in large numbers in mice with a congenital anaemia. The…

Investigation of Variant Haemoglobins and Thalassaemias

The haemoglobin molecule Human haemoglobin is formed from two pairs of globin chains each with a haem group attached. Seven different globin chains are synthesised in normal subjects; two, ε and ζ, are characteristic of the embryo and contribute to four transient embryonic haemoglobins referred to as haemoglobins Gower 1, Gower 2, Portland 1 and Portland 2. Haemoglobin F is the predominant haemoglobin of fetal life…

Acquired Haemolytic Anaemias

Assessing the likelihood of acquired haemolytic anaemia Haemolytic anaemia may be suspected from either clinical or laboratory abnormalities. Suggestive clinical features include anaemia, jaundice and splenomegaly. Other relevant clinical features that should be sought are a history of autoimmune disease, recent blood transfusion, recent infection, exposure to drugs or toxins, the presence of a cardiac prosthesis and risk of malaria. Previous clinical history and laboratory results…

Investigation of the Hereditary Haemolytic Anaemias : Membrane and Enzyme Abnormalities

The various initial steps to be taken in the investigation of a patient suspected of having a haemolytic anaemia are outlined in Chapter 11 and the changes in red cell morphology that may be found in haemolytic anaemias are illustrated in Chapter 5 . This chapter describes procedures useful in investigating haemolytic anaemias suspected to result from defects within the red cell membrane or deficiency of…

Laboratory Methods Used in the Investigation of the Haemolytic Anaemias

Red cells are typically removed from the circulation at the end of their lifespan of about 120 days. A shortened lifespan due to premature destruction may lead to haemolytic anaemia when bone marrow activity cannot compensate for the erythrocyte loss. The causes can be divided into three groups: 1. Defects within red cells from dysfunction of enzyme- controlled metabolism, abnormal haemoglobins or thalassaemias 2. Loss of…

Investigation of Megaloblastic Anaemia: Cobalamin, Folate and Metabolite Status

Acknowledgements Many thanks indeed to Dr Malcolm Hamilton and Mrs. Sheena Blackmore, authors of this chapter in editions 10 and 11 of this series. The descriptions of metabolic pathways for cobalamin, folate and homocysteine were assisted by Hematology Basic Principles and Practice and Homocysteine in Health and Disease. Thank you Annie Lee, Julie Bonser, staff at UK NEQAS Haematinics Department of Haematology and Good Hope NHS…

Iron Deficiency Anaemia and Iron Overload

Iron metabolism The iron content of the body and its distribution among the various proteins are summarised in Table 9-1 . Most of the iron is present in the oxygen-carrying protein of the red blood cell, haemoglobin, the synthesis and breakdown of which dominate iron turnover. Haem of haemoglobin is synthesised in nucleated red cells in the bone marrow and in reticulocytes by a pathway ending…

Molecular and Cytogenetic Analysis

Acknowledgements We would like to thank Dr Tom Vulliamy and other previous authors for their past contribution in the preparation of this chapter and we refer to past editions for a full extent of their work. Our understanding of the molecular basis of both inherited and acquired haematological disorders is now considerable and there are several ways in which this knowledge is being applied in diagnostic…

Bone Marrow Biopsy

Biopsy of the bone marrow is an indispensable adjunct to the study of diseases of the blood and may be the only way in which a correct diagnosis can be made. Marrow can be obtained by needle aspiration, percutaneous trephine biopsy or surgical biopsy. When performed correctly, bone marrow aspiration and trephine biopsy are simple and safe procedures that can be repeated many times and can…

Supplementary Techniques Including Blood Parasite Diagnosis

Tests for the acute-phase response Inflammatory response to tissue injury (i.e. the acute-phase response) includes alteration in serum protein concentration, especially increases in fibrinogen, haptoglobin, caeruloplasmin, immunoglobulins (Ig) and C-reactive protein (CRP), and a decrease in albumin. The changes occur in acute infection, during active phases of chronic inflammation, with malignancy, in acute tissue damage (e.g. following acute myocardial infarction) and with physical injury. Measurement of…

Blood Cell Morphology in Health and Disease

Examination of a fixed and stained blood film is an essential part of a haematological investigation, and it cannot be emphasised too strongly that, to obtain maximum information from the examination, the films must be well spread, well stained and examined systematically. Details of the recommended procedure for examination are given later in this chapter. The most important red cell abnormalities, as seen in fixed and…

Preparation and Staining Methods for Blood and Bone Marrow Films

Preparation of blood films on slides Blood films should be made on clean glass slides. Films made on coverglasses have negligible advantages and are unsuitable for modern laboratory practice. Films may be spread by hand or by means of an automated slide spreader, the latter being either a stand-alone instrument or a component of an automated blood cell counter. Manual method Blood films can be prepared…

Basic Haematological Techniques

It is possible to use manual, semiautomated or automated techniques to determine the various components of the full blood count (FBC). Manual techniques are generally low cost with regard to equipment and reagents but are labour intensive; automated techniques entail high capital costs but permit rapid performance of a large number of blood counts by a smaller number of laboratory workers. Automated techniques are more precise,…

Reference Ranges and Normal Values

A number of factors affect haematological values in apparently healthy individuals. As described in Chapter 1 , these include the technique and timing of blood collection, the transport and storage of specimens, the posture of the subject when the sample is taken, the prior physical activity and the degree of ambulation (e.g. whether the subject is confined to bed or not). Variation in the analytical methods…

Collection and Handling of Blood

Acknowledgement The author wishes to acknowledge the contribution of previous authors of this chapter – the late Corrine Jury, Yutaka Nagai and the late Noriyuki Tatsumi – and of Gareth Ellis, who reviewed the content of this chapter. Following an informed decision to analyse a blood sample, a specimen must be safely and correctly procured. It is essential to be aware that variation in this pre-analytical…

Models for point-of-care testing of critical care analytes

Introduction Point-of-care testing (POCT) is now common in many near-patient and critical care settings. For blood gas and electrolyte testing, this includes operating rooms (ORs), intensive care units (ICUs), cardiac catheterization labs (CCL), emergency departments (ED), and many primary care clinic settings. POCT has several distinct benefits, such as minimizing or eliminating specimen transportation and processing which have the added benefits of minimizing preanalytical effects and…

Quality control in blood gas and critical care testing

Routine daily quality control on blood gas instruments Most current blood gas/electrolyte/metabolite analyzers have onboard internal quality control (QC) systems that automatically monitor the stability and performance of reagents, electrodes, and electronics of the analyzer. These control systems make these “hybrid” analyzers well-suited to use in both laboratory and Point-of-Care Testing (POCT) areas. Two such analyzers use easily replaced multiuse packs that contain controls, calibrants, rinse…

Collection and handling of samples: effects on blood gases, Na, K, ionized Ca, Mg, lactate, and phosphate analyses

Sources of preanalytical errors in blood gas and electrolyte testing Introduction There are numerous sources of preanalytical errors in laboratory testing. Among these are the following: Wrong test ordered Wrong patient collected Error in sample collection, including improper patient preparation Errors during specimen transport Errors during specimen processing This chapter will address only the last three of these. As noted by Karon ( ) and Azman…

Lactate physiology and diagnostic evaluation

Introduction By 1900, many physicians had noted metabolic acidosis in patients who were critically ill. In 1925, Clausen identified increased lactate along with acidosis in these patients, which gave rise to the condition of “lactic acidosis,” although later work showed the lactate and acid were produced in entirely separate biochemical reactions, as described recently in a review ( ) . In 1964, Broder and Weil observed…