CLINICAL LAB SCIENEC

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ESSENTIALS OF CLINICAL LABORATORY SCIENCE
Absorbance (nm)
0.50
0.40
0.30
0.20
0.10
0.0
0.0 mg 0.5 mg 1.0 mg 1.5 mg 2.0 mg 2.5 mg
Concentration of standard (1.5 mg/dL)
FIGURE 9-13 Beer’s law graph.
Source: Delmar/Cengage Learning.
of the blood sample. The operation of a spectrophotometer
is indicated in Figure 9-14. As
shown by the line in Figure 9-13, a standard
with a concentration of 1.5 mg/dL gives a spectrophotometric
reading of 0.30. If an unknown
sample is read under the same conditions, and
the unknown sample yields a 0.42 absorbance,
calculations for the value of the unknown
would be as follows:
Absorbance of unknown/Absorbance of standard × Measured concentration of standard =
Value of unknown
0.42/0.30 × 1.5 mg/dL = 2.1 mg/dL
Kinetic Enzyme Calculations
For enzyme calculations, conditions are extremely important in determining a
factor for obtaining the value for an enzymatic reaction. A common enzyme is
alkaline phosphatase; bone tissue is rich with this enzyme. The reaction is a rate
reaction that should be linear as the enzyme reacts on the specific substrate. This
examination is also performed on a spectrophotometer, but instead of measuring
the color development as is done for some end-point determinations, this procedure
measures the rate of change for a specific period of time, which may vary
by instrumentation and by manufacturer of the reagent. The purpose of showing
this representative calculation for determining enzyme concentration in serum or
Spectrum
White light
Monochromatic light
Transmitted light
Electrical signal
0.276
Slit in
wavelength
detector
(D) PHOTO-DETECTOR
(E) DISPLAY
(A) LIGHT
SOURCE
(B) MONO-
CHROMATOR
(C) CUVETTE WITH
COLORED SOLUTION
FIGURE 9-14 How a spectrophotometer works, with diagram of internal parts.
Source: Delmar/Cengage Learning.
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