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Average the duplicate readings for each standard and sample and subtract the average zero standard optical density. Create a standard curve by reducing the data using computer software capable of genecanineing a four parameter logistic (4-PL) curve fit. As an alternative, construct a standard curve by plotting the mean absorbance for each standard on the x-axis against the concentcanineion on the y-axis and draw a best fit curve through the points on the graph. The data may be linearized by plotting the log of the target antigen concentcanineion versus the log of the O.D. and the best fit line can be determined by regression analysis. This procedure will produce an adequate but less precise fit of the data. If samples have been diluted, the concentcanineion read from the standard curve must be multiplied by the dilution factor.