TABLE 3.

Outline of the MaGiK method of TCR repertoire analysis

T-cell source	Spectratyping steps
Controls	1. Perform CDR3 spectratyping on the CD4+-T-cell subsets of ≥10 healthy controls. Fetal thymocytes or cord blood also make excellent controls but frequently are more difficult to obtain. Spectratyping on a capillary-based instrument such as the ABI model 310 Genetic Analyzer yields reproducible fluorescence intensity and fragment length data.
	2. Import the spectratyping data into a database application such as Microsoft Access.
	3. For each Vβ family, identify the discrete nucleotide ranges that comprise each CDR3 length.
	4. Compute the mean and standard deviation for each CDR3 length.
Test samples	1. Perform CDR3 spectratyping on the samples.
	2. Import the spectratyping data into a database application such as Microsoft Access.
	3. For each CDR3 length in a sample, compute a sample-to-control ratio by dividing the sample value by the control mean.
	4. Next, for each Vβ family, calculate the median sample-to-control ratio.
	5. Multiply each CDR3 length proportion in each Vβ family by its respective median sample-to-control ratio. This achieves a standardization of sample values based on a common control set.
	6. Establish and apply a criterion that defines clonal expansions. Values that exceed the control means by more than 3 standard deviations are statistically significant (P < 0.05). Biological significance may vary depending on the context, thus necessitating adjustment of the expansion definition in some instances.