MSI Analysis System, Version 1.2, Technical Manual

Instructions for Use of Product(s)

Literature # TM255

The MSI Analysis System, Version 1.2, is a fluorescent PCR-based assay to detect MSI in human cells. Typically, MSI analysis involves comparing allelic profiles of microsatellite markers generated by amplification of DNA from matching normal and test samples, which may be MMR deficient. Alleles that are present in the test sample but not found in the corresponding normal samples indicate MSI. The MSI Analysis System includes fluorescently labeled primers for co-amplification of seven markers including five mononucleotide repeat markers (BAT-25, BAT-26, NR-21, NR-24 and MONO-27) and two pentanucleotide repeat markers (Penta C and Penta D). The mononucleotide markers are used for MSI determination, and the pentanucleotide markers are used to detect potential sample mixups or contamination. An Internal lane size standard is added to the PCR samples to assure accurate sizing of alleles and to adjust for run-to-run variation. The PCR products are separated by capillary electrophoresis (CE), and the output data may be analyzed with fragment analysis software, such as GeneMapper® software (Applied Biosystems), to determine MSI status of test samples. To simplify data analysis, there are available panels and bins text files that automatically assign genotypes using GeneMapper® software.

Summary of Changes
The following changes were made to the 2/23 revision of this document:
1. Removed instructions for ABI PRISM® 310 Genetic Analyzer.
2. Updated Sections 1, 3, 4, 5, 7 and 9.
3. Provided instrument instructions in Section 6, including the Spectrum Compact CE Instrument.
4. Added additional thermal cycling protocols (Section 9.A).
5. Moved to a new template.

Revised 2/23.