nonlinear curve fitting -...

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Nonlinear Curve Fitting

Earl F. GlynnScientific Programmer

Bioinformatics

11 Oct 2006

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Nonlinear Curve Fitting• Mathematical Models• Nonlinear Curve Fitting Problems

– Mixture of Distributions– Quantitative Analysis of Electrophoresis Gels– Fluorescence Correlation Spectroscopy (FCS)– Fluorescence Recovery After Photobleaching (FRAP)

• Linear Curve Fitting• Nonlinear Curve Fitting

– Gaussian Case Study– Math– Algorithms– Software

• Analysis of Results– Goodness of Fit: R2

– Residuals• Summary

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Mathematical Models

• Want a mathematical model to describe observations based on the independent variable(s) under experimental control

• Need a good understanding of underlying biology, physics, chemistry of the problem to choose the right model

• Use Curve Fitting to “connect” observed data to a mathematical model

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Nonlinear Curve Fitting ProblemsMixture Distribution Problem

Adapted from www.math.mcmaster.ca/peter/mix/demex/expike.html

0 20 40 60 80

0.00

0.01

0.02

0.03

0.04

0.05

0.06

Heming Lake Pike: Length Distribution

Length [cm]

Pro

babi

lity

Den

sity

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Nonlinear Curve Fitting ProblemsMixture Distribution Problem

Data are fitted by five normal distributions with constant coefficient of variation

0 20 40 60 80

0.00

0.01

0.02

0.03

0.04

0.05

0.06

Heming Lake Pike: Distribution by Age Groups

Length [cm]

Pro

babi

lity

Den

sity

ex

xf σ

µ

σπ2

2

2)(

221)(

−−

=

Normal Probability Density Function

Coefficient of Variation

µσ

=cv

6

Deconvolve a pixel profile of a banding pattern into a family of Gaussian or Lorentzian curves

Quantitative Analysis of Electrophoresis Gels

Takamato, et al, Nucleic Acids Research, 32(15), 2004, p. 2

Nonlinear Curve Fitting Problems

http://papakilo.icmb.utexas.edu/cshl-2005/lectures/CSHL_Lecture05_khodursky.ppt#23

Das, et al, RNA (2005), 11:348

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Nonlinear Curve Fitting ProblemsQuantitative Analysis of Electrophoresis Gels

Many proposed functional forms besides Gaussian or Lorentzian curves

DiMarco and Bombi, Mathematical functions for the representation of chromatographic peaks,Journal of Chromatography A, 931(2001), 1-30.

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Nonlinear Curve Fitting ProblemsFluorescence Correlation Spectroscopy (FCS)

Bacia, Kim & Schwille, “Fluorescence cross-correlation spectroscopy in living cells,”Nature Methods, Vol 3, No 2, p. 86, Feb. 2006.

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Nonlinear Curve Fitting ProblemsFluorescence Correlation Spectroscopy (FCS)

From discussion by Joe Huff at Winfried Wiegraebe’s Lab Meeting, 11 Aug 2006

Note likely heteroscedasticity in data

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Nonlinear Curve Fitting ProblemsFluorescence Recovery After Photobleaching (FRAP)

From discussion by Juntao Gao at Rong Li’s Lab Meeting, 25 Sept 2006

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Nonlinear Curve Fitting ProblemsFluorescence Recovery After Photobleaching (FRAP)

From discussion by Juntao Gao at Rong Li’s Lab Meeting, 25 Sept 2006

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Linear Curve Fitting

• Linear regression• Polynomial regression• Multiple regression• Stepwise regression• Logarithm transformation

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Given data points ( , ).

We want the “best” straight line, ( , ),through these points, where is the“fitted”value at point :

xixi

yi

Linear Regression: Least Squares

yiˆ

xy ii ba +=ˆ

yiˆ


14

xixi


xy ii ba +=ˆ

0 1 2 3 4

01

23

45

(x,y) Data

x

y


Linear Fit

Error Function

[ ]∑ ⋅+−=

=N

iii xbayba

1

22 )(),(χ

(xi,yi)

Assume homoscedasticity (same variance)

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Search (a,b) parameter space to minimize error function, χ2


b

0

1

2

3

a

-1 0 1 2

50

100

150

200

χ2 Error Function

[ ]∑ ⋅+−=

=N

iii xbayba

1

22 )(),(χ

Linear Fit

9.1)9.0,2.1(2

=χ

xy ii 9.02.1ˆ +=


16

xixi

Linear Regression: Least SquaresLinear Curve Fitting

0 1 2 3 4

01

23

45

Least Squares Line

x

yy = 1.2 + 0.9x

How can (a,b) parameters be found directly without a search?

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xixi


• Differentiate χ2 with respect to parameters a and b• Set derivatives to 0.


How can (a,b) parameters be found directly without a search?

021

2

=⋅−−−=∂

∂ ∑=

N

iii xbay

aχ

0)(21

2

=⋅−−−=∂

∂ ∑=

N

iii i xbayx

bχ

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xixi

Linear Regression: Least SquaresHow can (a,b) parameters be found directly without a search?

xy ii ba +=ˆLinear Fit Simultaneous Linear Equations


=

∑∑

∑∑∑

yxy

ba

xxxN

ii

i

ii

i2

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xixi

Linear Regression: Least SquaresHow can (a,b) parameters be found directly without a search?

xy ii ba +=ˆLinear Fit


=

∑∑

∑∑∑

yxy

ba

xxxN

ii

i

ii

i2

i x y x² xy1 0 1 0 02 1 3 1 33 2 2 4 44 3 4 9 125 4 5 16 20

Sum 10 15 30 39

=

3915

3010105

ba

2.15060

101030510393015

301010530391015

==⋅−⋅⋅−⋅

==a

9.05045

10103051510395

30101053910155

==⋅−⋅⋅−⋅

==b

Simultaneous Linear Equations

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xixi


> x <- 0:4> y <- c(1,3,2,4,5) > summary( lm(y ~ x) )

Call:lm(formula = y ~ x)

Residuals:1 2 3 4 5

-0.2 0.9 -1.0 0.1 0.2

Coefficients:Estimate Std. Error t value Pr(>|t|)

(Intercept) 1.2000 0.6164 1.947 0.1468 x 0.9000 0.2517 3.576 0.0374 *---Signif. codes: 0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1

Residual standard error: 0.7958 on 3 degrees of freedomMultiple R-Squared: 0.81, Adjusted R-squared: 0.7467F-statistic: 12.79 on 1 and 3 DF, p-value: 0.03739

R solutionusing lm (linear model)

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xixi


Assume homoscedasticity (σi = constant = 1)

Assume heteroscedasticity

∑

⋅+−

=

=N

i

ii

i

xbayba

1

2

2 )(),(

σχ

Often weights σi are assumed to be 1.Experimental measurement errors can be used if known.

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ex

xf σ

µ

σπ2

2

2)(

221)(

−−

=Normal Probability Density Function

Gaussian Case Study

-2 -1 0 1 2 3 4 5

0.0

0.1

0.2

0.3

0.4

0.5

0.6

Gaussian Data

x

y

x y-2.0 0.00004-1.5 0.00055-1.0 0.00472-0.5 0.027390.0 0.107480.5 0.285391.0 0.512751.5 0.623352.0 0.512752.5 0.285393.0 0.107483.5 0.027394.0 0.004724.5 0.000555.0 0.00004

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ex

xf σ

µ

σπ2

2

2)(

221)(

−−

= [ ]∑ −=

=N

ii xify

1

22

)(),( σµχ

mu

-50

5

sigma

0.4

0.6

0.8

1.0

1.2

1.4

2

4

6

8

χ2

Gaussian Case Study

Minimumμ = 1.5σ = 0.8

Gradient descentworks well onlyinside “valley”here

Assume homoscedasticity

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Nonlinear Curve FittingGaussian Case Study

Derivatives may be useful for estimating parameters

-4 -2 0 2 4

0.0

0.1

0.2

0.3

0.4

Single Gaussian

x

y

-4 -2 0 2 4

-0.2

0.0

0.2

1st Derivative

x

y'

-4 -2 0 2 4

-0.4

-0.2

0.0

0.2

2nd Derivative

x

y''

U:/efg/lab/R/MixturesOfDistributions/SingleGaussian.R

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Nonlinear Curve FittingGaussian Case Study

Derivatives may be useful for determining number of terms

0 20 40 60 80

0.00

0.02

0.04

0.06

x

y

0 20 40 60 80

-0.0

050.

005

1st Derivative

x

y'

0 20 40 60 80

-0.0

06-0

.002

0.00

2

2nd Derivative

x

y''

Heming Lake Pike

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Nonlinear Curve FittingGiven data points (xi,yi).Given desired model to fit (not always known):

y = y(x; a)where there are M unknown parameters:

ak, k = 1,2,..., M.The error function (“merit function”) is

∑

−

=

=N

i

ii

i

axyya

1

2

2 );()(

σχ

From Press, et al, Numerical Recipes in C (2nd Ed), 1992, p. 682

Math

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Need to search multidimensional parameter space to minimize error function, χ2

Nonlinear Curve FittingMath

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Gradient of χ2 with respect to parameters a will be zero at the minimum:

MkN

i k

i

i

ii

k aaxyaxyy

a,...,2,1

12

2 );(]);([2 =∑

= ∂∂−

−=∂∂

σχ


Taking the second derivative of χ2:

From Press, et al, Numerical Recipes in C (2nd Ed), 1992, p. 682

∑=

∂∂

∂−−∂

∂∂

∂=

∂∂∂ N

i k

iii

i

k

i

ik aaaxyaxyy

aaxy

aaxy

aa 1

2

2

22 );(]);([

);();(12lll σ

χ

αkl = Hessian or “curvature” matrix (after dropping “2”)

Often small and ignored

βk (after dropping “-2”)

Math

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• Levenberg-Marquardt is most widely used algorithm:– When “far” from minimum, use gradient descent:

– When “close” to minimum, switch to inverse Hessian:

• “Full Newton-type” methods keep dropped term in second derivative – considered more robust but more complicated

• Simplex is an alternative algorithm

βα k

M

k a =∆∑=

l

l

l

1

β ll ⋅=∆ constanta

Nonlinear Curve FittingAlgorithms

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Nonlinear Curve Fitting• Fitting procedure is iterative• Usually need “good” initial guess, based on

understanding of selected model• No guarantee of convergence• No guarantee of optimal answer• Solution requires derivatives: numeric or

analytic can be used by some packages

Algorithms

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Nonlinear Curve FittingSoftware

IDL: curvefit function; MPFIT: Robust non-linear least square curve fitting(3 limited licenses)• Joe Huff in Advanced Instrumentation is quite-well versed in using MPFIT and applying it in IDL

R: nls function• many statistics• symbolic derivatives (if desired)• flawed implementation: exact “toy” problems fail unless “noise” added

MatLab: Curve Fitting Toolbox(1 limited license)

Mathematica(1 limited license)

PeakFit: Nonlinear curve fitting for spectroscopy, chromatography and electrophoresis(1 limited license)

OriginPro: Peak Fitting Module(10 limited licenses)

32http://www.itl.nist.gov/div898/strd/general/dataarchive.html

NIST reference datasets with certified computational results

Nonlinear Curve FittingSoftware

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Analysis of Results• Goodness of Fit: R2

• Residuals

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Goodness of Fit: R2Coefficient of Determination

Percentage of Variance Explained

Mean] for [Corrected (SS) Squares of Sum Total(RSS)Squares of Sum Residual

−= 12R

( )( )∑ −

∑ −−=

yyyy

Ri

ii2

2

2 ˆ1 10 2 ≤≤ R

• “Adjusted” R2 compensates for higher R2 as terms added. • A “good” value of R2 depends on the application.• In biological and social sciences with weakly correlated

variables, and considerable noise, R2 ~ 0.6 might be considered good.

• In physical sciences in controlled experiments, R2 ~ 0.6 might be considered low.

Faraway, Linear Models with R, 2005, p.16-18

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Residuals

“Curve fitting made easy,” Marko Ledvij, The Industrial Physicist, April/May 2003. http://www.aip.org/tip/INPHFA/vol-9/iss-2/p24.html

• Residuals are estimates of the true and unobservable errors.• Residuals are not independent (they sum to 0).

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Analysis of Residuals

• Are residuals random?• Is mathematical model appropriate?• Is mathematical model sufficient to

characterize the experimental data?• Subtle behavior in residuals may suggest

significant overlooked propertyGood Reference: “Analysis of Residuals: Criteria for Determining Goodness-of-Fit,”Straume and Johnson, Methods in Enzymology, Vol. 210, 87-105, 1992.

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Analysis of ResidualsSynthetic FRAP Data: Fit with 1 term when 2 terms are better

Near “perfect” fit, but why is there a pattern in the residuals?

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Analysis of ResidualsLomb-Scargle periodogram can indicate “periodicity” in the residuals

Flat line with all “bad” p-values would indicate “random” residuals

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Analysis of ResidualsSynthetic FRAP Data: Fit with 2 terms

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Analysis of ResidualsFCS Data and Heteroscedasticity

Heteroscedasticity in Residuals

∑

−

=

=N

i

ii

i

axyya

1

2

2 );()(

σχ

Scaled Residuals

Scaling Factor

FCS Residual Plots Courtesy of Joseph Huff, Advanced Instrumentation & Physics

Use F Test to test for unequal variances

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Analysis of ResidualsHeteroscedasticity and Studentized Residuals

See http://en.wikipedia.org/wiki/Studentized_residual

• Studentized residual is a residual divided by an estimate of its standard deviation

• The “leverage” hii is the ith diagonal entry of a “hat matrix.”

hiii

i

−=

1ˆˆ

σεResidualdStudentize

• Externally Studentized Residuals follow Student’s t-distribution.

• Can be used to statistically reject “outliers”

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Summary• A mathematical model may or may not be appropriate for

any given dataset.• Linear curve fitting is deterministic.• Nonlinear curve fitting is non-deterministic, involves

searching a huge parameter space, and may not converge.• Nonlinear curve fitting is powerful

(when the technique works).• The R2 and adjusted R2 statistics provide easy to

understand dimensionless values to assess goodness of fit.• Always study residuals to see if there may be unexplained

patterns and missing terms in a model.• Beware of heteroscedasticity in your data. Make sure

analysis doesn’t assume homoscedasticity if your data are not.

• Use F Test to compare the fits of two equations.

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AcknowledgementsAdvanced Instrumentation & Physics• Joseph Huff• Winfried Wiegraebe

nonlinear curve fitting -...

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