Handling Data and Figures of Merit

Data comes in different formats

time
Histograms
Lists
But.
Can contain the same information about quality
What is meant by quality?
(figures of merit)
Precision, separation (selectivity), limits of detection,
Linear range
day weight day weight day weight
1 140 31 143.9 61 144
2 140.1 32 144 62 144.2
3 139.8 33 142.5 63 144.5
4 140.6 34 142.9 64 144.2
5 140 35 142.8 65 143.9
6 139.8 36 143.9 66 144.2
7 139.6 37 144 67 144.5
8 140 38 144.8 68 144.3
9 140.8 39 143.9 69 144.2
10 139.7 40 144.5 70 144.9
11 140.2 41 143.9 71 144
12 141.7 42 144 72 143.8
13 141.9 43 144.2 73 144
14 141.4 44 143.8 74 143.8
15 142.3 45 143.5 75 144
16 142.3 46 143.8 76 144.5
17 141.9 47 143.2 77 143.7
18 142.1 48 143.5 78 143.9
19 142.5 49 143.6 79 144
20 142.3 50 143.4 80 144.2
21 142.1 51 143.9 81 144
22 142.5 52 143.6 82 144.4
23 143.5 53 144 83 143.8
24 143 54 143.8 84 144.1
25 143.2 55 143.6
26 143 56 143.8
27 143.4 57 144
28 143.5 58 144.2
29 142.7 59 144
30 143.7 60 143.9
My weight
Plot as a function of time data was acquired:
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Day
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Do not use curved lines to connect data
points
that assumes you know more about the
relationship of the data than you really do
Comments:
background is white (less ink);
Font size is larger than Excel
default (use 14 or 16)
day weight day weight day weight
1 140 31 143.9 61 144
2 140.1 32 144 62 144.2
3 139.8 33 142.5 63 144.5
4 140.6 34 142.9 64 144.2
5 140 35 142.8 65 143.9
6 139.8 36 143.9 66 144.2
7 139.6 37 144 67 144.5
8 140 38 144.8 68 144.3
9 140.8 39 143.9 69 144.2
10 139.7 40 144.5 70 144.9
11 140.2 41 143.9 71 144
12 141.7 42 144 72 143.8
13 141.9 43 144.2 73 144
14 141.4 44 143.8 74 143.8
15 142.3 45 143.5 75 144
16 142.3 46 143.8 76 144.5
17 141.9 47 143.2 77 143.7
18 142.1 48 143.5 78 143.9
19 142.5 49 143.6 79 144
20 142.3 50 143.4 80 144.2
21 142.1 51 143.9 81 144
22 142.5 52 143.6 82 144.4
23 143.5 53 144 83 143.8
24 143 54 143.8 84 144.1
25 143.2 55 143.6
26 143 56 143.8
27 143.4 57 144
28 143.5 58 144.2
29 142.7 59 144
30 143.7 60 143.9
Bin refers to what groups of
weight to cluster. Like
A grade curve which lists
number of students who got
between 95 and 100 pts
95-100 would be a bin
Assume my weight is a single, random, set of similar data
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Weight (lbs)
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Make a frequency chart (histogram) of the data
Create a model of my weight and determine average
Weight and how consistent my weight is
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Weight (lbs)
#

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s
= measure of the consistency, or similarity, of weights
average
143.11
s = 1.4 lbs
Inflection pt
s = standard deviation
Characteristics of the Model Population
(Random, Normal)
Peak height, A
Peak location (mean or average),
Peak width, W, at baseline
Peak width at half height, W
1/2
Standard deviation, s, estimates the variation in an infinite population, o
Related concepts
( )
f x
A
e
x
=


|
\

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.
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o t

o
2
1
2
2
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
0.4
0.45
-5 -4 -3 -2 -1 0 1 2 3 4 5
s
A
m
p
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Width is measured
At inflection point =
s
W
1/2
Triangulated peak: Base width is 2s < W < 4s
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
0.4
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-5 -4 -3 -2 -1 0 1 2 3 4 5
s
A
m
p
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+/- 1s
Area +/- 2s = 95.4%
Area +/- 3s = 99.74 %
pp s ~ 6
Pp = peak to peak or
largest separation of
measurements
Peak to peak is sometimes
Easier to see on the data vs time plot
Area = 68.3%
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Peak to
peak
pp s ~ 6
139.5
144.9
s~ pp/6 = (144.9-139.5)/6~0.9
(Calculated s= 1.4)
0
5
10
15
20
25
Weight (lbs)
#

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s
-0.05
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
0.4
0.45
-5 -4 -3 -2 -1 0 1 2 3 4 5
s
A
m
p
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Scale up the first derivative and second derivative to see better
There are some other important characteristics of a normal (random)
population
1
st
derivative
2
nd
derivative
-1
-0.8
-0.6
-0.4
-0.2
0
0.2
0.4
0.6
-5 -4 -3 -2 -1 0 1 2 3 4 5
s
A
m
p
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Population, 0
th
derivative
1
st
derivative,
Peak is at the inflection
Determines the std. dev.
2
nd
derivative
Peak is at the inflection
Of first derivative should
Be symmetrical for normal
Population; goes to zero at
Std. dev.
Asymmetry can be determined from principle component analysis
A. F. (Alanah Fitch) = asymmetric factor
Is there a difference between my baseline weight and school weight?
Can you detect a difference? Can you quantitate a difference?
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Day
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Vacation
School Begins
Baseline
Comparing TWO populations of measurements
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138 139 140 141 142 143 144 145 146 147
Weight (lbs)
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Exact same information displayed differently, but now we divide
The data into different measurement populations
baseline
school
Model of the data as two normal populations
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138 139 140 141 142 143 144 145 146 147
Weight (lbs)
#

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s
Average
Baseline weight
Average school
weight
Standard deviation
Of baseline weight
Standard deviation
Of the school weight
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Weight (lbs)
#

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s
0
5
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138 139 140 141 142 143 144 145 146 147
Weight (lbs)
#

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We have two models to describe the population of measurements
Of my weight.
In one we assume that all measurements fall into a single population.
In the second we assume that the measurements
Have sampled two different populations.
Which is the better model?
How to we quantify better?
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138 139 140 141 142 143 144 145 146 147
Weight (lbs)
#

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0
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138 139 140 141 142 143 144 145 146 147
Weight (lbs)
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Compare how close
The measured data
Fits the model
Did I gain weight?
The red bars represent the difference
Between the two population model and
The data
The purple lines represent
The difference between
The single population
Model and the data
Which model
Has less summed
differences?
This process (summing of the squares of the differences)
Is essentially what occurs in an ANOVA

Analysis of variance
Normally sum the square of the difference in order to account for
Both positive and negative differences.
In the bad old days you had to work out all the sums of squares.
In the good new days you can ask Excel program to do it for you.
Anova: Single Factor
5% certainty
SUMMARY
Groups Count Sum Average Variance
Column 1 12 277.41 23.1175 8.70360227
Column 2 12 345.72 28.81 6.50010909
ANOVA
Source of Variation SS df MS F P-value F crit
Between Groups 194.4273 1 194.4273 25.5762995 4.59E-05 4.300949
Within Groups 167.2408 22 7.601856 Source of Variation
Total 361.6682 23
Test: is F<F
critical
? If true = hypothesis true, single population
if false = hypothesis false, can not be explained
by a single population at the
5% certainty level
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
14 19 24 29 34 39
Length (cm)
F
r
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c
y
White, N=12, Sum sq diff=0.037, stdev=2.55
White, N=38, Sum sq diff=0.028, stdev=2.15
Red, N=12, Sum sq diff=0.11, stdev=3.27
Red, N=40, Sum sq diff=0.017, stdev-2.67
0
0.05
0.1
0.15
0.2
0.25
0.3
14 19 24 29 34 39
Length, cm
F
r
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q
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c
y
N=24 Sum sq diff=0.0449, stdev=3.96
N=78, sum sq diff=0.108, stdev=4.05
In an Analysis of Variance you test the hypothesis that the sample is
Best described as a single population.
1. Create the expected frequency (Gaussian from normal error curve)
2. Measure the deviation between the histogram point and the expected
frequency
3. Square to remove signs
4. SS = sum squares
5. Compare to expected SS which scales with population size
6. If larger than expected then can not explain deviations assuming a
single population
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
14 19 24 29 34 39
Length (cm)
F
r
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c
y
White, N=12, Sum sq diff=0.037, stdev=2.55
White, N=38, Sum sq diff=0.028, stdev=2.15
Red, N=12, Sum sq diff=0.11, stdev=3.27
Red, N=40, Sum sq diff=0.017, stdev-2.67
0
0.05
0.1
0.15
0.2
0.25
0.3
14 19 24 29 34 39
Length, cm
F
r
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q
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c
y
N=24 Sum sq diff=0.0449, stdev=3.96
N=78, sum sq diff=0.108, stdev=4.05
0
0.005
0.01
0.015
0.02
0.025
0.03
0.035
0.04
15 17 19 21 23 25 27 29 31 33 35
Length (cm)
S
q
u
a
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e

D
i
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f
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c
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E
x
p
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d

M
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d
The square differences
For an assumption of
A single population
Is larger than for
The assumption of
Two individual
populations
There are other measurements which describe the two populations
Resolution of two peaks
R
x x
W W
a b
a b
=

+
2 2
Mean or average
Baseline width
0
0.5
1
1.5
2
2.5
3
3.5
4
4.5
1 1.5 2 2.5 3 3.5 4
x
S
i
g
n
a
l
x
a
x
b
x x
a b

W
a
2
W
b
2
In this example
W W
a b
2 2
+
Peaks are baseline resolved when R > 1
R x x
W W
a b
a b
> > + 1
2 2
:
0
0.2
0.4
0.6
0.8
1
1.2
1.4
1.6
1.8
1 1.5 2 2.5 3 3.5 4
x
S
i
g
n
a
l
x
a
x
b
x x
a b

W
a
2
W
b
2
In this example
W W
a b
2 2
+
Peaks are just baseline
resolved when R = 1
R x x
W W
a b
a b
= = + 1
2 2
:
0
0.2
0.4
0.6
0.8
1
1.2
1.4
1.6
1 1.5 2 2.5 3 3.5 4
x
S
i
g
n
a
l
x
a
x
b
x x
a b

W
a
2
W
b
2
In this example
W W
a b
2 2
+
Peaks are not baseline resolved
when R < 1
R x x
W W
a b
a b
< < + 1
2 2
:
2008 Data
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
14 19 24 29 34 39
Length (cm)
F
r
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q
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c
y
White, N=12, Sum sq diff=0.037
Red, N=12, Sum sq diff=0.11
What is the R for this data?
( )
Ax W W
p R W
< +
1
2
R < 1
Visually less resolved
Visually better resolved
Comparison of 1978 Low Lead to 1979 High Lead
0
5
10
15
20
25
0 20 40 60 80 100 120 140 160
Seri es2 Seri es3
Comparison of 1978 Low Lead to 1978 High Lead
0
5
10
15
20
25
0 20 40 60 80 100 120 140 160
IQ Verbal
%

M
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d
Anonymous 2009 student analysis of Needleman data
( )
( )
W
W
a
b
2
112 70 42
2
130 95 35
= =
= =
~ ~
~ ~
R
x x
W W
a b
a b
=

+
2 2
Visually less resolved
Visually better resolved
Comparison of 1978 Low Lead to 1979 High Lead
0
5
10
15
20
25
0 20 40 60 80 100 120 140 160
Seri es2 Seri es3
Comparison of 1978 Low Lead to 1978 High Lead
0
5
10
15
20
25
0 20 40 60 80 100 120 140 160
IQ Verbal
%

M
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Anonymous 2009 student analysis of Needleman data
( )
( )
W
W
a
b
2
112 70 42
2
130 95 35
= =
= =
~ ~
~ ~
( )
x x a b = = ~ ~ 112 95 17
R
x x
W W
a b
a b
=

+
+
=
2 2
17
42 35
022 ~ .
Other measures of the quality of separation of the
Peaks
1. Limit of detection
2. Limit of quantification
3. Signal to noise (S/N)
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
0.4
0.45
-6 -4 -2 0 2 4 6 8 10 12
s
A
m
p
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0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
0.4
0.45
-6 -4 -2 0 2 4 6 8 10 12
s
A
m
p
l
i
t
u
d
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3s
X
blank
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
0.4
0.45
-6 -4 -2 0 2 4 6 8 10 12
s
A
m
p
l
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t
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3s
X
limit of detection
x x s
LOD blank blank
= + 3
99.74%
Of the observations
Of the blank will lie
below the mean of the
First detectable signal
(LOD)
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
0.4
0.45
-6 -4 -2 0 2 4 6 8 10 12
s
A
m
p
l
i
t
u
d
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3s
Two peaks are visible when all the data is summed together
Estimate the LOD (signal) of this data
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Day
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0
5
10
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138 139 140 141 142 143 144 145 146 147
Weight (lbs)
#
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Other measures of the quality of separation of the
Peaks
1. Limit of detection
2. Limit of quantification
3. Signal to noise (S/N)
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
0.4
0.45
-6 -4 -2 0 2 4 6 8 10 12
s
A
m
p
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x x s
LOQ blank blank
= + 9
Your book suggests 10
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
0.4
0.45
-6 -4 -2 0 2 4 6 8 10 12
s
A
m
p
l
i
t
u
d
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9s
Limit of quantification requires absolute
Certainty that no blank is part of the
measurement

0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
0.4
0.45
-6 -4 -2 0 2 4 6 8 10 12
s
A
m
p
l
i
t
u
d
e
139
140
141
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143
144
145
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0 10 20 30 40 50 60
Day
w
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(
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b
s
)
Estimate the LOQ (signal) of this data
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143
144
145
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0 10 20 30 40 50 60
Day
w
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(
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0
5
10
15
20
25
138 139 140 141 142 143 144 145 146 147
Weight (lbs)
#
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s
Other measures of the quality of separation of the
Peaks
1. Limit of detection
2. Limit of quantification
3. Signal to noise (S/N)
Signal = x
sample
- x
blank
Noise = N = standard deviation, s
S
N
x x
s
x x
pp
sample
blank
sample
blank
=

~

|
\

|
.
|
6
Estimate the S/N of this data
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Day
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(
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Vacation
School Begins
Baseline
Signal
Peak to peak variation within
mean school
~ 6s where s = N for Noise
(This assumes pp school ~ pp baseline)
0
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15
20
25
138 139 140 141 142 143 144 145 146 147
Weight (lbs)
#
o
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s
0
5
10
15
20
25
30
35
0 5 10 15 20 25 30
Sample number
l
e
n
g
t
h

(
c
m
)
Can you tell where the switch between
Red and white potatoes begins?
What is the signal (length of white)?
What is the background (length of red)?
What is the S/N ?
Effect of sample size on the measurement
Error curve
Peak height grows with # of measurements.
+ - 1 s always has same proportion of total number of measurements
However, the actual value of s decreases as population grows
s
s
n
sample
population
sample
=
22.5
23
23.5
24
24.5
25
25.5
26
26.5
27
0 2 4 6 8 10 12 14
Sample number
R
e
d

R
u
n
n
i
n
g

L
e
n
g
t
h

A
v
e
r
a
g
e
0
0.5
1
1.5
2
2.5
3
3.5
4
4.5
5
R
e
d

R
u
n
n
i
n
g

S
t
d
e
v

2008 Data
y = -0.8807x + 5.9303
R
2
= 0.9491
2.5
2.7
2.9
3.1
3.3
3.5
3.7
3.9
4.1
1.5 2 2.5 3 3.5 4
sqrt number of samples
s
t
d
e
v

r
e
d

l
e
n
g
t
h

c
m
s
s
n
sample
population
sample
=
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
14 19 24 29 34 39
Length (cm)
F
r
e
q
u
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n
c
y
White, N=12, Sum sq diff=0.037, stdev=2.55
White, N=38, Sum sq diff=0.028, stdev=2.15
Red, N=12, Sum sq diff=0.11, stdev=3.27
Red, N=40, Sum sq diff=0.017, stdev-2.67
Calibration Curve
A calibration curve is based on a selected measurement as linear
In response to the concentration of the analyte.
Or a prediction of measurement due to some change
Can we predict my weight change if I had spent a longer time on
Vacation?
bx a y + =
( ) vacation on days b a lbs fitch + =
0
5
10
15
20
25
138 139 140 141 142 143 144 145 146 147
Weight (lbs)
#

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b
s
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a
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s
( ) vacation on days b a lbs fitch + =
5 days
The calibration curve contains information about the sampling
Of the population
y = 0.3542x + 140.04
R
2
= 0.7425
139
139.5
140
140.5
141
141.5
142
142.5
143
0 1 2 3 4 5 6
Days on Vacation
F
i
t
c
h

W
e
i
g
h
t
,

l
b
s
Can get this by using trend line
y = -0.8807x + 5.9303
R
2
= 0.9491
2.5
2.7
2.9
3.1
3.3
3.5
3.7
3.9
4.1
1.5 2 2.5 3 3.5 4
sqrt number of samples
s
t
d
e
v

r
e
d

l
e
n
g
t
h

c
m
This is just a trendline
From format data
Sample sqrt(#samples) stdev
1 1 #DIV/0!
2 1.414213562 2.036468
3 1.732050808 4.475727
4 2 4.31441
5 2.236067977 3.844045
6 2.449489743 3.844604
7 2.645751311 3.735124
8 2.828427125 3.458414
9 3 3.235055
10 3.16227766 3.093053
11 3.31662479 2.935944
12 3.464101615 2.950187
SUMMARY OUTPUT
Regression Statistics
Multiple R 0.296113395
R Square 0.087683143
Adjusted R Square -0.013685397
Standard Error 0.703143388
Observations 11
ANOVA
df SS MS F Significance F
Regression 1 0.427662048 0.427662 0.864994 0.376617
Residual 9 4.449695616 0.494411
Total 10 4.877357664
Coefficients Standard Error t Stat P-value Lower 95%
Intercept 3.884015711 0.514960076 7.542363 3.53E-05 2.719094
X Variable 1 -0.06235252 0.067042092 -0.93005 0.376617 -0.21401
Using the analysis
Data pack
Get an error
Associated with
The intercept
In the best of all worlds you should have a series of blanks
That determine youre the noise associated with the background
x x s
LOD blank blank
= + 3
Sometimes you forget, so to fall back and punt, estimate
The standard deviation of the blank from the linear regression
But remember, in doing this you are acknowledging
A failure to plan ahead in your analysis
x x b conc LOD
LOD blank
= + [ . ]
[ . ] conc LOD
s
b
blank
=
3
Extrapolation of the associated error
Can be obtained from the Linear
Regression data
Sensitivity (slope)
x x s
LOD blank blank
= + 3
x s x b conc LOD
blank
blank
blank
+ = + 3 [ . ]
The concentration LOD depends on BOTH
Stdev of blank and sensitivity
Signal LOD
!!Note!!
Signal LOD Conc LOD
We want Conc. LOD
-350
-300
-250
-200
-150
-100
-50
0
0 2 4 6 8 10 12
pH or pM
m
V
y = -31.143x - 74.333
R
2
= 0.9994
-350
-300
-250
-200
-150
-100
-50
0
0 2 4 6 8 10 12
pH or pM
m
V
y = -31.143x - 74.333
R
2
= 0.9994
-350
-300
-250
-200
-150
-100
-50
0
0 2 4 6 8 10 12
pH or pM
m
V
y = -31.143x - 74.333
R
2
= 0.9994
y = -41x - 118.5
R
2
= 0.9872
-350
-300
-250
-200
-150
-100
-50
0
0 2 4 6 8 10 12
pH or pM
m
V
Difference in slope is one measure selectivity
In a perfect method the sensing device would have zero
Slope for the interfering species
Selectivity
Pb
2+
H
+
Limit of linearity
5% deviation
Summary: Figures of Merit Thus far
R = resolution
S/N
LOD = both signal and concentration
LOQ
LOL
Sensitivity (calibration curve slope)
Selectivity (essentially difference in slopes)
Can be expressed in terms of signal, but better
Expression is in terms of concentration
Tests: Anova
Why is the limit of detection important?
Why has the limit of detection changed so much in the
Last 20 years?
The End
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Which of these two data sets would be likely
To have better numerical value for the
Ability to distinguish between two different
Populations?
Needlemans data
2008 Data
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
14 19 24 29 34 39
Length (cm)
F
r
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q
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c
y
White, N=12, Sum sq diff=0.037
Red, N=12, Sum sq diff=0.11
Height for normalized
Bell curve <1
Which population is more variable?
How can you tell?
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
14 19 24 29 34 39
Length (cm)
F
r
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q
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n
c
y
White, N=12, Sum sq diff=0.037, stdev=2.55
White, N=38, Sum sq diff=0.028, stdev=2.15
Red, N=12, Sum sq diff=0.11, stdev=3.27
Red, N=40, Sum sq diff=0.017, stdev-2.67
Increasing the sample size decreases the std dev and increases separation
Of the populations, notice that the means also change, will do so until
We have a reasonable sample of the population
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