The TCR BV repertoire analyzed by CDR3 size profiles.

PBMC cDNA was used as template in the primary BV-BC PCR reaction. Subsequently, a secondary, “run-off” PCR reaction with an internal, FAM-labeled BC primer was performed. The products were run on an acrylamide gel and displayed through the use of an automated sequencer. Immunoscope software analysis reveals the distribution of TCR BV mRNA sizes. Within each peak are represented mRNAs of the same length (horizontal axis); the vertical axis represents relative fluorescence intensity. The CDR3 size profiles of the 24 TCR BV families of a healthy individual and of a patient with PNH are shown. Numbers in the right upper corner indicate the corresponding BV family. A diverse (normal) repertoire consists of 6 to 9 peaks distanced from each other by exactly 3 nucleotides (1 codon), and distributed in a Gaussian or nearly Gaussian fashion. As determined by the 3-criteria analysis (see “Patients, materials, and methods”), in this healthy individual, TCR family BV18 is skewed. In the patient with PNH, families BV3, BV7, BV9, BV14, BV16, BV20, BV21, and BV24 are skewed.