Three Microsoft Excel calibration programs for the laboratory

This logit regression program in Excel is optimised to work with RIA and ELISA-data.
The program can be used for fitting all sigmoid and half-sigmoid curves and even a straight line.
Logit, 4-parameter or Rodbard regression

The PLA presented here is written in Microsoft Excel and is an copy of the method described by D.J. Finney.
Up to thirteen samples can be calculated
by hand, from an ELISA plate reading or list of responses.
Parallel line assay PLA

The SRA presented here is written in Microsoft Excel and is an copy of the method described by D.J. Finney.
Slope ratio assay SRA

See below what is the 'best' calibration program to use.


HP41-C programs

I started learning programming on a HP-9715 desktop calculator at my laboratory in the end of the 1970's.
When the HP41C handheld calculator arrived on the market the same programming RPN-language was used and programming continued at home.
I made several programs and send them to the HP "Users' program library Europe" so they could be used by everyone.
Unfortunately this library is lost. I retained a copy of most of my listings.
These have been
scanned and they are available for reference and their formulas.

If time comes I may add more. More in The museum of HP calculators

Listing of files

What calibration program is the best to use?

SRA, PLA or Logit?

In following example the data from an ELISA was fed into the three methods.
The calibration line is entered in the logit regression program.
The data pairs that were suitable to be entered in the SRA are marked purple and the dose-responses that could be used in the PLA are coloured red in the picture below.
For the SRA and PLA only those parts that are linear after transformation can be used resulting is a smaller measuring range that can be entered.

Logit,PLA or SRA

SRA

PLa

The results for the sample were calculated in:
PLA as 265 10%
SRA as 274 10%
Logit as 261 8%

My conclusions:

As to be expected the calculation method has no great influence on the result.
All results fall well within in the  confidence limits of the three methods.


Benefits Logit
The measuring range is large.
The rejection on SD of calculated result is based on historical data.
Cons Logit
Does not test directly whether sample dilutes parallel with the standard.
Because of the long measuring range one should be careful with the hook effect or prozone effect.
NB both cons will be noted because the average will have a bad standard deviation and be flagged)

Benefits PLA
Robust accepted calculation method with descriptive statistics.
Disadvantages PLA
Relatively small measuring area.
The statistics is based on F-tests; when a standard has perfect duplicates all sample results becomes invalid. In practice this results is a rejection of a complete test so now and then because the calibration line was so perfect by accident. And also the other way round. Bad standard lines give valid results with large confidence intervals.

SRA benefits
Suitable for low concentrations and activity measurements.
Cons SRA
Often very small measuring range because the dose is linear.
The statistics is based on F-tests and not on historical performances of the test.

Final conclusion
My choice to use logit regession as first option is its simplicity and long measuring range. It is simple in its rejection criteria and it is easier to use in LIMS systems.
SRA and PLA are a better choice if linearity is an issue.

 

November 2009 - 2019

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Ed Nieuwenhuys Email