preparing analytical samples. For example, prior to
measuring the concentration of some protein-bound
and intracellular analytes, it is necessary to release
the analyte into solution. To accomplish this, a
release reaction will be performed on the test speci-
mens. The same reaction may not be performed on
the calibrators because the analyte in calibrators is
already in solution.
Precision of measurement
Precision is defined as the closeness of agree-
ment among the result values obtained from a large
number of replicate measurements (Dybkaer 1995).
Precision is measured on an ordinal scale but its
inverse, imprecision, is a quantitative measure.
Imprecision (or random error of measurement) is the
dispersion of results for a large number of replicate
measurements. It is usually expressed in terms of
standard deviations.
Imprecision arises from multiple sources which
can be categorized according to the following
scheme (Dybkaer 1995): (1) those that arise during a
single batch of measurements, (2) those that arise
over the course of the performance of multiple
batches of measurement, and (3) those that arise
when several laboratories contribute to the produc-
tion of results.
Imprecision arising during a single batch of
measurements is called within-run, or within-batch
imprecision and is measured in terms of the within-
run standard deviation. Because it quantifies
method precision in the setting of the minimum
number of sources of measurement variability,
within-run imprecision is the minimum precision
attainable by the method. The causes of within-run
imprecision include volumetric errors, instrumental
fluctuations, variability in the efficiency of the
separation step, and vagaries in the rate and
completeness of the signal generating step.
Imprecision arising from the performance of
multiple batches of measurement is called between-
run, or between-batch, imprecision and is measured
in terms of the between-run standard deviation. The
causes of between-run imprecision include recalibra-
tion of the measurement system, different calibrators
and reagents, different operators, and time
Laboratory Methods
2-5
Table 2.2
Selected Separation Techniques for Improving Method Specificity
Technique
Principle and examples
Membrane permeability analyte-permeable membrane separates sample from site of signal generation
only specific analyte passes through membrane
O
2
and CO
2
electrodes, ion-selective electrodes, ultrafiltration, equilibrium dialysis
Electrophoresis
chemicals in buffer migrate through support medium in electric field
chemicals have different migration rates due to different charges and different
degrees of interaction with stationary phase
specific analyte migrates to characteristic location
cellulose acetate electrophoresis, agarose gel electrophoresis (named for support medium)
Chromatography
mobile phase (gas or liquid) passes through a column with a stationary phase bound
to support medium
chemicals in mobile phase have different degrees of interaction with stationary phase
which leads to different migration rates through column
specific analyte elutes from column at characteristic retention time
gas chromatography, liquid chromatography (named for mobile phase)
Antibody binding
analyte-specific antibodies are bound to support medium
only specific analyte binds to antibodies and is retained on support medium
following wash
heterogeneous radio-, enzyme, and fluorescence immunoassays
