As the final step of the ELISA, the calculation of the test results is important for the entire experiment.

Take the sandwich method as an example.

1. The ELISA sample is usually set up with 1~2 duplicate wells for the average value of the absorbance of the standard and experimental samples. The measured value of a single sample should not exceed 20% of the average.

2. The average OD (absorbance) of the sample is subtracted from the OD average of the standard concentration of 0.

3. Establish a standard curve

A four-parameter logistic function curve is fitted on the logarithmic graph (standard concentration on the X-axis and corresponding OD value on the Y-axis). It is recommended to use the origin software.

The detailed steps are as follows:

Step 1 : Enter OD value and standard concentration

Step 2: Select the type of fit curve

Select the value box you entered and click the option as follows.

Step 3: Check "find specific X/Y"

After completing the above operation, click the Fit button, the fitting curve and related information will be automatically generated (as shown in the fourth step)

Step 4: View standard curve information (such as parameter values, R-Square, etc.)

Step 5: Set the axis type to logarithm.

Double-click the fitted curve in step 4 to edit it. Double-click the X-axis and Y-axis to set the coordinate type to "Log10".

Step 6: Calculate sample concentration based on the standard curve

Select the fourth worksheet (Fit NLF Find X from Y1) and enter the OD value of the sample in the left column. The corresponding concentration will be automatically generated in the right column.

If the OD value of the sample exceeds the upper limit of the standard curve, properly dilute the sample to obtain accurate results before the experiment. When analyzing the data, the actual concentration of the sample should be the concentration obtained on the standard curve multiplied by the dilution factor.

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