Clonality of CD3 negative large granular lymphocyte proliferations determined by PCR based X-inactivation studies.
OBJECTIVE
To examine persistent CD3-large granular lymphocytosis (LGL) cases for clonality, both by lineage specific (T cell receptor) and lineage independent (X-inactivation) molecular methods; and to find out whether X-inactivation studies are more appropriate than gene rearrangement studies for this subset of LGL disorders.


METHODS
Patients were selected who had LGL of more than six months' duration and identified as CD3- by immunophenotyping. T cell receptor studies and, where possible, X-inactivation studies of the phosphoglycerate kinase (PGK) gene were carried out. Analysis of subpopulations was carried out on cases heterozygous for PGK by the use of a polymerase chain reaction (PCR) method for X-inactivation.


RESULTS
Of 17 CD3- LGL cases studied, all were found to be germline for beta, gamma, and delta T cell receptor studies, and immunoglobulin heavy chain genes. However, six of these were analysed by X-inactivation of the PGK gene and two cases gave clonal band patterns but only within the CD3- subpopulation.


CONCLUSIONS
Clonal analysis by the lineage independent method of X-inactivation allows clonal expansion undetected by T and B cell specific markers to be identified. It is therefore a more appropriate method for the analysis of CD3- LGL. This has implications for diagnosis in CD3- LGL disorders.
Molecular Pathology and Haematological Malignancy Diagnostic Service, University of Leeds.
A
A
Kelly
S J
SJ
Richards
M
M
Sivakumaran
C
C
Shiach
A D
AD
Stewart
B E
BE
Roberts
C S
CS
Scott
D000328:Adult; D000368:Aged; D001483:Base Sequence; D017252:CD3 Complex / Q000097:blood*; D002999:Clone Cells / Q000276:immunology; D004303:Dosage Compensation, Genetic; D005260:Female; D015321:Gene Rearrangement; D015329:Gene Rearrangement, T-Lymphocyte; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000235:genetics; D018189:Immunomagnetic Separation; D016130:Immunophenotyping / Q000379:methods; D007694:Killer Cells, Natural / Q000276:immunology*; D008218:Lymphocytosis / Q000201:enzymology / Q000235:genetics* / Q000276:immunology; D008875:Middle Aged; D008969:Molecular Sequence Data; D010735:Phosphoglycerate Kinase / Q000235:genetics; D016133:Polymerase Chain Reaction
D017252:CD3 Complex; D007143:Immunoglobulin Heavy Chains; D010735:Phosphoglycerate Kinase