Biochemistry and hematology results have a role in detection of clinical or subclinical disease, diagnosis, monitoring of illness, and confirming health.1–3 The determined values have no meaning unless there is some basis for comparison so they can be interpreted as either “normal” or “not normal.”² Typically, this interpretation is made by comparing the result obtained with a population-based reference interval that describes the central 95% of a healthy population.⁴ This means that, by definition, some healthy individuals (2.5% below and 2.5% above the ­reference limits) will fall outside the reference interval, with increasing confidence that results at a greater distance outside the reference limits represent abnormality.

The generation of such reference intervals requires careful selection of the individual reference animals to ensure they are healthy and that the age, sex, breed, reproductive status, and other factors (e.g., diet, vaccination, and deworming status) are representative of the population to which this type of reference interval will be applied. There are extensive recommendations for numbers of reference individuals and statistical techniques to be used.⁵

Caution should be applied to accepting reference intervals from the literature or those provided by instrument manufacturers, or even those provided by commercial pathology laboratories since these may not be applicable to the patient population of all veterinary practices. One study found that less than 30% of biochemistry reference intervals provided from multiple sources to be appropriate.⁶ Further, reference intervals are not transferrable across different analyzers or methods.⁶ To avoid misclassification of patients based on provided reference intervals, clinicians should consider determining reference intervals for the population of their practice or conducting reference interval transference validation in which 20 samples from healthy animals are checked against the provided reference intervals (Box 9.1).^6–8

A newer paradigm for interpretation of results is to compare results from an individual to prior results when that subject was known to be well.2,9 Such a comparison can only be made if the inherent physiologic fluctuation of the parameter being measured is known; this inherent fluctuation is known as biological variation. Biological variation data have only recently become available for cats,^9–11 and are used to determine a reference change value (RCV), a percentage change, above or below which is indicative of a statistically significant change.² The RCV can then be applied to the mean of an individual cat’s results when healthy to determine an individualized reference interval that looks like traditional, population reference intervals with upper and lower limits in each analyte’s usual units¹² but can only be used for the cat it was determined for (Box 9.2).

Population-based reference intervals have limited diagnostic sensitivity when variation between individuals is wider than variation within the individuals comprising the population. The applicability of population reference intervals is determined by a ratio of variation that occurs within individuals to variation that occurs across a group. Individualized reference intervals can be used in all cases.12 As with population-based reference intervals, comparisons can only be made when the results being assessed are generated with the same analyzer and using the same method.^2,12–15

Currently, few laboratories flag changes in serial results according to probability of statistical significance based on biologic variation. As the use of biological variation information becomes more widespread in veterinary medicine, the authors envision that this will become the standard for reporting on serial changes in results to make clinical decisions for the best possible patient care.

Each laboratory result represents a range of possible values, and a given laboratory result is impacted not just by the presence or absence of disease, but also by biological variation of the measurement in question and analytical variation of the equipment used to make the measurement. This range of possible values is known as dispersion and can be calculated.16–18 For simplicity, clinical pathology is usually taught in terms of discrete results, as will be used for the rest of this chapter. However, it is important to recognize that because a discrete result represents a range, results that are just within a reference interval (whether population-based or individual-based) may actually represent an increase and, conversely, results just outside a reference interval may be normal for the individual patient.¹² As with individualized reference intervals, the authors have an expectation that dispersion will be included in standard reporting of clinical pathology results in the not too distant future.