What Supplies To Line Up Ahead Of Your ELISA Assay Development and Validation?

Author : mariabrownema
Publish Date : 2021-04-07 05:15:52
 What Supplies To Line Up Ahead Of Your ELISA Assay Development and Validation?

Enzyme-linked immunosorbent assay (ELISA) is a bioanalytical immunoassay widely used in the drug development process. It has been continuously developed and modified for over 50 years, with novel technologies significantly upgrading its usefulness as a routine technique in medical and clinical practices. Hence, these immunoassays must be capable of providing valid and reliable study results.

During immunoassay development and validation, optimal and accurate performance is the final goal of an immunoassay protocol. An ELISA assay can be used routinely, only after it has proven to provide precise, accurate, and reproducible results, achieved through a standardized protocol. Hence, ELISA assay development and validation becomes important if scientists want to use it in clinical and nonclinical diagnostics. Let us understand what supplies laboratories should line up before moving on with ELISA development and validation.

What are the supplies needed for ELISA development and validation?

ELISA development begins with a concrete immobilization strategy. Thus, the first requirement before beginning assay development is ELISA well plates for the solid phase of an immunoassay method. Generally, a 96 or 384 well plate made up of polyvinyl/polystyrene is used in ELISA assays. The next part of the immobilization strategy involves beads. Beads help attach the molecule of interest to the surface of ELISA wells. Once the immobilization strategy is in place, the samples are prepared for the experiment. Additional steps and resources may be needed to purify the samples.

Antigen and antibody interactions govern the whole ELISA assay protocol. A highly pure antigen efficiently captures an antibody, and hence, an appropriate antigen is the most crucial element of an ELISA development and validation. The next component is a reporter enzyme. Enzyme antibody conjugation is the key for signal output observed in an ELISA. Alkaline phosphatase and horseradish peroxidase are commonly used reporter enzymes. These enzymes are conjugated to several proteins and antibodies for maximum activity retention. The final step of an ELISA is the detection step. The antigen is detected using an enzyme-substrate unit. An enzyme-substrate complement produces a color whose intensity is directly proportional to the bound antigen in ELISA plates.

Ideally, all the supplies are complete in the development phase itself. However, quality control samples are another component required in Elisa assay validation. Quality control samples are needed to test the specificity of an ELISA assay. They are samples with known concentrations prepared in a matrix similar to the sample of interest. If the results obtained from quality control samples are satisfactory, only then the method is deemed valid. Similar to specificity, the accuracy of the ELISA method is evaluated using quality control samples. Hence, quality control samples are crucial in validating several assay characteristics.

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Conclusion

ELISA is one of the most widely used analytical techniques for detecting and quantifying drugs and drug products. ELISA is based on antigen-antibody interaction, and all the types of ELISA protocols are developed keeping these interactions in the picture. Only after careful optimization, development, and validation of assay characteristics, a reliable and robust immunoassay method becomes available for routine diagnostic and biomedicine use.



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