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ELISA Overview

Note: Although this software and accompanying documentation is dated 2004-2005, it is still valid in 2014. Questions can be sent to CDC-INFO.

ELISA for Windows is a series of programs or program modules which process bioassay data collected from 96 well ELISA plates downloaded from several different models of ELISA readers. If a user's reader is not represented in the program, instructions are provided as to how to create a simple ASCII text data file for import into the program. ELISA for Windows then performs a series of analyses on the processed data. This software is fully validated and the validation documents are available for download.

Briefly, the 96-well ELISA plate is formatted by placing a sequence of diluted standard sera in a set of contiguous wells on the plate (vertical or horizontal configuration). The program offers a flexible template design module so that plates may be formatted differently for different assays. Serum samples and quality control specimens are then placed in the remaining wells of the plate. Serum samples may be either single dilutions or a sequential dilution series. All standards, patient unknowns and quality control samples may be replicated one or more times, within each dilution, to increase the precision of resulting calculations. An ELISA plate reader collects optical density measurements from each well and the operator imports these absorbance values to a desktop computer and stores the data as an ASCII text file. ELISA for Windows is able to abstract the standard series, individual serum samples, and quality control samples from this file. The standards data are used to form a characteristic or standard curve which may be modeled using a three point cubic spline or a four parameter logistic-log function. The four parameters of the logistic function may be estimated using two methods: iteratively reweighted least squares and robust procedures. Estimation options include the Taylor series linearization (Gauss-Newton) and the Marquardt's compromise estimation algorithms. The standard curve is then used to interpolate antibody concentrations for the patient isolates and quality control samples. Summary statistics are calculated from these concentrations (means, standard deviations, coefficients of variation, etc). ELISA for Windows also forms plots of the standards data with the estimated standard curve superimposed on the data points.

These programs are in the public domain and are compiled in 32-bit mode using Microsoft Visual Basic. Interested users are welcome to download the documentation and programs and test them in their laboratories with the proviso that each investigator will need to spend some time learning the functions of each module to benefit from the whole collection of programs. The authors also request that they be cited as a source for this work when these techniques are used in the preparation of published reports. A suggested citation for ELISA for Windows appears at the bottom of the title page of the user's manual.

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