Specimen

Whole Blood EDTA

Collection Medium:

Minimum:

Preferred minimum:  3 mL whole blood EDTA
Adults and Peds absolute minimum: 2 mL whole blood EDTA

Rejection Criteria:

Rejected if specimen is greater than 72 hours from blood draw, frozen 
or clotted whole blood or bone marrow samples.

Analytic Time:

2 weeks

Reference Range:

The BCR-ABL allele was amplified from CML patient's plasma sample in 
the first round one step RT-PCR. The ABL kinase domain was amplified in 
the semi-nested PCR followed by direct sequencing using ABI/prism 
Big-Dye terminator cycle sequencing kit on automated capillary DNA 
sequencer (ABI Prism(R) 3100 Genetic Analyzer). Changes of nucleotides 
in the coding sequence as well as amino acid will be reported.

The sensitivity of the RT-PCR is one leukemia cell in the background of 
100,000 normal cells. The sensitivity of detecting mutant allele is 20% 
in the background of wild-type cells. The sequencing encompasses ABL 
exon 4 to exon 9 (ATP binding domain and activation loop). This assay 
detects only mutations and/or polymorphisms that are within the 
amplicon (863 bp) generated. Mutations that lie outside the boundaries 
of the regions amplified will not be detected. Sequence changes that 
are in the PCR primer sites will also not be detected. Furthermore, 
sequence polymorphisms in the PCR primer sites, if exists in patient's 
DNA being analyzed, may result in failure to amplify the associated 
allele, therefore, may potentially result in failure to detect the 
mutation that is associated with that allele..

Comments:

The plasma based molecular assay is based on the new concept that in 
hematological diseases, tumor cell pour into their circulation their 
DNA and RNA, these components can be detected in plasma. Imatinib 
mesylate (ST1571;Gleevec) is a selective BCR-ABL kinase inhibitor, 
effective in the treatment of chronic myelogenous leukemia (CML). Most 
patients in chronic phase maintain durable responses; however, many in 
blast crisis fail to respond, or relapse quickly. ABL kinase domain 
mutations are the most commonly identified mechanism associated with 
relapse. The molecular monitoring in the first few months of therapy 
may play a crucial role in detecting patients at high risk of Imatinib 
resistance.  This ABL kinase mutation assay may detect drug-resistant 
mutations before clinical relapse and identify candidate suitable for 
alternative therapy.

Methodology:

Reverse Transcriptase Polymerase Chain Reaction, Nested PCR, Sequencing

CPT Code:

83891
83894
83898x6
83902
83904
83909
83912