LAUSR

Dear Editor, Bias in hemolysis analyte results due to an increase in the analyte concentration caused by intracellular release (e.g., potassium, phosphate, magnesium, AST, ALT, lactate dehydrogenase, folate, or urea) or a decrease in the concentration because of dilution by intracellular contents (e.g. sodium, chloride, or albumin) is often significant before spectrophotometric or chemical interference by Hb or other intracellular contents is manifested [1]. Therefore, the hemolysate preparation method selected for interference studies must provide an accurate representation of the real analyte changes due to hemolysis. The osmotic shock hemolysate preparation procedure, originally developed to study analytical interference from dissolved Hb [2], is commonly used in interference studies. In a popular adaptation of this procedure [3], after three saline washes, the red blood cells are hemolyzed by adding an equal volume of deionized water (DIW) followed by a freeze–thaw cycle. The obtained hemolysate is added to serum to prepare samples with a desired Hb concentration. In addition to changes due to hemolysis, the sample composition is altered by the substitution of the serum with DIW in the added hemolysate; therefore, the analyte result of the test pool is compared with that of a control prepared by adding an equal volume of saline to the same serum pool [3]. Most other hemolysate preparation methods, e.g., methods that use multiple freeze–thaw cycles or shear stress hemolysis, do not alter the sample composition beyond changes due to hemolysis, and the result of the test pool is compared with that of a non-hemolyzed base pool [4]. The scenario of intracellular release of an analyte, using a modeled sample with a serum urea concentration of 6.0 mmol/L, is illustrated in Fig. 1A. The concentration of urea in red blood cells is approximately 1.2 times the concentration in serum and was considered 7.2 mmol/L in this case [5]. Modeled analyte concentrations in samples prepared for paired interference testing using the osmotic shock hemolysate preparation procedure were compared with those in a sample prepared by the freeze–thaw or mechanical hemolysis method. As the sample composition does not change beyond changes due to hemolysis in the latter two methods, equivalence of the final analyte concentrations was assumed, and the methods are presented together. As equal volumes of washed red cells and DIW are used in the osmotic shock procedure, for simplicity of comparison, Hb, 150 g/L; hematocrit, 50%; and complete hemolysis were assumed. Contributions from white blood cells and platelets were ignored to simplify the explanation. In the prepared sample, the urea concentration increased by 7% compared to that in the control in the osmotic shock method and by 1.0% compared to that in the base pool in the other two methods. The difference in the scenario of dilution by cellular contents, taking sodium as an example, is illustrated in Fig. 1B [4]. For sodium (and chloride), it is preferred to compare the result with a control prepared by adding an equal volume of DIW rather