The demonstration of intravascular haemolysis by means of serum paper electrophoresis and a modification of Schumm's reaction.

In studying the pathogenesis of anaemia associated with many clinical disorders one is often confronted with the problem of whether intravascular haemolysis is taking place. Haemolysis may result either from an intrinsic abnormality of the red cell, rendering it of shorter life span than normal, and this may be of an hereditary or acquired nature, or, alternatively, from the presence of some plasma factor capable of inducing lysis. Plasma factors producing lysis result from certain types of poisoning, toxic effects of drugs, and many generalized infections. Haemolysis may also occur as a result of renal or hepatic failure and in other metabolic disorders. The exact mechanism of the haemolysis in these conditions, although unknown, is probably also attributable to plasma factors. Despite the wide range of known causative factors many cases of haemolysis remain unclassified, e.g., those which are seen in the reticuloses and other neoplastic states. In patients manifesting an extreme degree of haemolysis, extracorpuscular haemoglobin can be seen in the serum, which, if in sufficient concentration, is excreted into the urine. More commonly an increased rate of haemolysis is manifest by a raised serum bilirubin level and an increased excretion of urobilinogen in the urine and faeces. Reticulocytosis, if present in such circumstances, reflects a compensatory hyperplasia of the bone marrow, but both these latter diagnostic features may be partially obscured should the rate of bilirubin production be matched by the ability of the liver to excrete it, or should some complicating factor, such as uraemia or chronic infection, simultaneously depress erythropoietic activity. Under these conditions even moderate haemolysis may escape detection by existing routine laboratory procedures, and in the present work two new complementary techniques are described which appear to be more sensitive than other simple procedures, and can demonstrate that haemolysis is occurring or has recently taken place. These tests depend on showing an altered pattern of haemoglobin attachment to the serum proteins. One technique employs paper electrophoresis of serum with subsequent differential staining of the strips to show the position of the protein bands relative to the haemoglobin pigments, and the other technique is based on a variation of the Schumm haemochromogen reaction (Schumm, 1912), in which the haemoglobin itself is involved. The Schumm reaction, as normally performed, allows detection of methaemalbumin in much lower concentration than is possible by direct spectroscopic examination (Fairley, 1941). The presence of methaemalbumin in serum is indicative of intravascular haemolysis, of which the occasional occurrence in pernicious anaemia argues for a significant haemolytic component in that disease. The common finding of raised serum bilirubin levels in patients with the disease points to a similar conclusion, and it appeared possible that closer examination of such sera and specimens from other patients in whom haemolysis was occurring or suspected might yield information concerning the severity of the haemolytic process, and its response to therapy in relation to the patients' haematological and clinical progress.