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Linearity
1
Linear range
Range of concentra.ons where the
signals are directly propor1onal to the
concentra1on of the analyte
in the sample.
Sensi.vity
The change in instrument response
which corresponds to a change in
the measured quan1ty.
The gradient of the response curve.
2
Experiment planning
for evalua.on of linearity
• Type of calibra1on samples
• Concentra1on range
• Number of concentra1on levels
• Measurement protocol
• Matrix-containing samples
– Matrix-matched calibra1on
– Blank matrix extract (preferably the same type as the sample)
3
Concentra.on range
• Appropriate for the method
• Expected working range ± 10%…20%
• 70%…130% of expected analyte concentra1on
• Blank sample
4
Measurement protocol
• As similar as possible to the real life situa1on
– Random order
– Between the samples
Evalua.on of linearity
• Visual evalua.on
• Residuals
5
Visual evalua.on
120000000 80000000
LINEAR 70000000 NOT LINEAR
100000000
60000000
80000000
50000000
Signal (au)
Signal (au)
60000000 40000000
30000000
40000000
20000000
20000000
10000000
0 0
0 2 4 6 8 10 0 1 2 3 4 5 6
c (mg/kg) c (mg/kg)
Residual
• Difference between experimental signal and calculated signal
yi ŷi
120000000
100000000
yi
80000000
Signal (au)
60000000
0
0 2 4 6 8 10 εi = yi − ŷi
c (mg/kg)
6
Visual evalua.on
120000000 140000000
80000000
LINEAR 70000000
120000000
NOT LINEAR
LINEAR ?
100000000
60000000
100000000
80000000
Signal (au)
Signal (au)
50000000
80000000
Signal
60000000 40000000
60000000
30000000
40000000
40000000
20000000
20000000 20000000
10000000
0 0 0
0 2 4 6 8 10 0 0 1 2 2 4 6
3 8
4 10
5 12
6
c (mg/kg) c (mg/kg)
c (mg/kg)
Plot of residuals
7
Linear, homoscedas.c
10000000
8000000
6000000
Residual (au)
4000000
2000000
0
0 0.2 0.4 0.6 0.8 1 1.2
-2000000
-4000000
-6000000
c (mg/kg)
Linear, heteroscedas.c
30000000
20000000
10000000
Residual (au)
0
0 2 4 6 8 10 12
-10000000
-20000000
-30000000
c (mg/kg)
8
Not linear, heteroscedas.c
30000000
20000000
10000000
Residual (au)
0
0 2 4 6 8 10 12
-10000000
-20000000
-30000000
-40000000
c (mg/kg)
Rela.ve residuals
• ±20% is acceptable
yi − ŷi
Yi =
ŷi
9
Correla.on coefficient r
Coefficient of determina.on r2
• Total variability in the response that is
accounted for by the model
Linear range
10
“range”
120000000
100000000
Signal (au)
80000000
60000000
40000000
20000000
0
0 2 4 6 8 10 12
c (mg/kg)
11
Dynamic range, Analy1cal range
Working range, Calibra.on range
120000000
100000000
Signal (au)
80000000
60000000
40000000
*LOQ
20000000
0
0 2 4 6 8 10 12
c (mg/kg)
80000000
60000000
40000000
*LOQ
20000000
0
0 2 4 6 8 10 12
c (mg/kg)
12
Sensi.vity
• gradient of the calibra1on graph
Sensi.vity
120,000,000 ΔSignal
Slope =
100,000,000 Signal = 7610033⋅ c + 32270418 Δc
80,000,000
Signal (au)
ΔSignal
60,000,000
40,000,000
20,000,000 Δc
0
0 2 4 6 8 10
c (mg/kg)
13
Sensi.vity
• Method op1miza1on
• Quality assurance
Sy. x
LoQ = 10 ×
Slope
14