Conventional endpoint PCR requires that the PCR product be fully amplified before detection. At the end of the PCR cycles, the product is analyzed to determine if the target DNA sequence was amplified. These results are often obtained by performing agarose gel electrophoresis. Gel electrophoresis separates all the PCR products by size. These results are then compared against a gel with DNA fragments of known size and molecular weight.
There are many limitations to the endpoint analysis of standard PCR, including:
- The detection procedures are very time consuming.
- Determination based on size is not completely accurate.
- Agarose gel detection has low resolution and sensitivity, is only semi quantitative, and cannot be automated.