The study of submicroscopic or minimal residual disease (MRD) in childhood acute lymphoblastic leukemia may eventually lead to stratification of therapy on an individual patient basis (reviewed in ref. 1 ). PCR of immunoglobulin heavy chain (IgH) and T-cell receptor (TCR) gene rearrangements provides widely informative markers (Table 1 ), which, in the majority of cases, are stable during the disease course (2 ). Generation of leukemia-specific probes using this technique allows detection of MRD at levels of one leukemic cell in 10,000 to 100,000 normal bone marrow mononuclear cells (BM MNC). Table 1Primer Systemsa
|
IgH FR3 |
1 |
5′-ACACGGC(C/T)(G/C)TGTATTACTGT-3′(2) |
3 |
5′-GTGACCAGGGT(C/T)C C(C/T)TGGCCCCAG-3′ (2) |
1.5–30 |
65–155 |
75% |
8% |
|
4 |
5′-AACTGCAGAGGAGACGGTGACC-3′(3) |
15–30 |
80–170 |
|||||
|
2 |
5′-GACCAGGGT(C/T)C C(C/T)TGGCCCCAG-3′e |
|||||||
|
Vδ2-Dδ3 |
5 |
5′-CTTGCACCATCAGAGAGAGA-3′(2) |
7 |
5′-GTTTTTGTACAGGTCTCTGT-3′ |
10–30 |
100–150 |
45% |
4% |
|
8 |
5′-AGGGAAATGCACTTTTGCC-3′(2) |
15–30 |
110–170 |
|||||
|
6 |
5′-TTTTGTACAGGTCTCTGT-3′c |
|||||||
|
Vδ1-Jδ1 |
5′-GCCTTACAGCTAGAAGATTC-3′ |
5′-GTTCCTTTTCCAAAGATGAG-3′ |
15–30 |
80–150 |
5% |
25% |
||
|
VγI-Jγ1/2d |
5′-TG(A/C)(C/T)TCTGG(A/G)GTCTATTACTGT-3′ |
5′-CGATACTTACCTGTGACAAC(C/A)AG-3′ |
30 |
80–160 |
45% |
90%d |
||
|
VγII-Jγ-1/2d |
5′-AAACAGGACATAGCTACCTACT-3′ |
5′-CGATACTTACCTGTGACAAACC/AAG-3′ |
30 |
80–160 |
45% |
90%d |
||
|
Lead Vγ2-anti Vγ2 |
5′-GTCATGTCAGCCATTGAGTT-3′ |
5′-TCTCTCTCTGATGGTGCAAG-3′ |
15 |
220 |
control |
control |
aThe primer shown here use a common buffer and generate products that can easily be resolved on 8% PAGEbRefers to Fig. 1cSequencing primersdThese primers can be used in a multiplex reaction. The VγI primer is a consensus primer and amplifies all members of this group except Vγ7 + B and +T indicate the percentage of patients by lineage expected to show a clonal rearangement at each locus at diagnosis.
