Advanced Precision Medicine Laboratory - Clinical Services

• FFPE Slides- uncoated unbaked slides in plastic slide containers
  • 1 H&E slide and 10 adjacent unstained 5um sections. Area of tumor cell content should be a minimum of 3x3mm or 5,000 cells and be comprised of at least 70% cancer cells. Necrosis and inflammation in the area of the highest neoplastic content should be mild or less.
• FFPE Block- send in sealed biohazard bag
  • Area of tumor cell content should be a minimum of 3x3mm or 5,000 cells and be comprised of at least 70% cancer cells. Necrosis and inflammation in the area of the highest neoplastic content should be mild or less.
• DNA:
  • Sample Concentration: ≥ 12.5 ng/uL
  • Sample Quality: < 5 (▲CT)
  • Amount of Material: ≥ 250 ng
• Oncology Test Requisition

Samples that don't meet the above acceptance criteria may fail to generate useable data.

• Turnaround Time: 10-14 days from sample receipt
• JAX OncoMethyl Array Sample Report

The JAX OncoMethyl™ Array utilizes a machine learning algorithm for the classification of central nervous system (CNS) tumors based on genomic methylation profiling data. The JAX OncoMethyl™ Array uses genomic DNA extracted from FFPE tissues (≥70% neoplastic content) that is followed by bisulfite conversion (Zymo Research). Converted DNA undergoes whole genome amplification and is processed utilizing the Infinium MethylationEPIC Array (Illumina). Raw IDAT files are processed through the CNS methylation classifier developed by the Molecular Neuropathology group at the German Cancer Research Center (DKFZ) (PMID: 29539639). Methylation Class Family and Methylation Class calibrated scores are provided by the classifier. During validation, 98% of samples with Methylation Class calibrated scores ≥0.84 were considered “classifiable” and resulted in either confirmation, refinement, or reassignment of diagnosis.

• FFPE Slides- uncoated unbaked slides in plastic slide containers
  • 1 H&E slide and 10 adjacent unstained 5um sections. Area of tumor cell content should be a minimum of 3x3mm or 5,000 cells and be comprised of at least 30% cancer cells.
• FFPE Block- send in sealed biohazard bag
  • Area of tumor cell content should be a minimum of 3x3mm or 5,000 cells and be comprised of at least 30% cancer cells.
• DNA:
  • Sample Concentration: ≥ 10 ng/uL
  • Sample Quality: < 5 (▲CT)
  • Amount of Material: ≥ 200 ng
• Oncology Test Requisition

Samples that don't meet the above acceptance criteria may fail to generate useable data.

• Turnaround Time: 5-7 days from sample receipt
• JAX MGMT Promoter Methylation Sample Report

The JAX MGMT Promoter Methylation Assay utilizes a quantitative PCR (qPCR) followed by high-resolution melt analysis (HRM) to identify MGMT promoter methylation. Genomic DNA is extracted from FFPE tissues (minimum 30% neoplastic content) and bisulfite treated using the EZ DNA Methylation-Gold Kit (Zymo). The bisulfite-treated DNA is amplified via qPCR followed by a melting analysis on an Applied Biosystems QuantStudio 7. The area under the curve (AUC) is calculated for the HRM derivative plots for both methylated and unmethylated peaks and the ratio of methylated to unmethylated is calculated. Specimen will be interpreted as MGMT Promoter Unmethylated if the methylated/unmethylated ratio falls within the validated unmethylated range. Specimen with ratios above a 15% methylated control will be interpreted as MGMT Promoter Methylated and specimen with ratios between unmethylated and 15% will be interpreted as Indeterminate.

• Formalin-fixed paraffin embedded (FFPE) specimens, including unstained cut slide specimens are acceptable.
  • Decalcified specimens are not accepted.
  • Purified nucleic acids are not accepted.
  • Surface Area: minimum of 3x3mm tumor area.
  • Neoplastic Content: Minimum 30% neoplastic content within the tumor area.
• Oncology Test Requisition

Samples that don't meet the above acceptance criteria may fail to generate useable data.

• Turnaround Time: 14-21 days from sample receipt
• JAX SOMASEQ™ Automated Sample Report

JAX SOMASEQ™ uses genomic DNA and RNA extracted from macro dissection-enriched FFPE tissue sections (≥30% neoplastic content), followed by enrichment of target exons and introns by hybrid-capture (Illumina). Illumina sequencers generate 101bp paired-end sequence reads with a median exon coverage of greater than or equal to 150X. Variants within regions that do not meet our coverage thresholds are not reported. For a list of these regions, please contact cgl_cs@jax.org. The LOD (limit of detection) for SNVs and indels was determined as 5%. The LOD for CNVs was 5 copies for amplifications and 1 copy for deletions. Mutational analysis is performed using the TSO500 bioinformatic pipeline. Variants were called against human genome build GRCh37.

Testing of affected and/or unaffected family members can aide in the interpretation of variants identified on the JAX Genome test. Testing options for family members include:

• JAX Comparator Genome Sequencing: Comparator sequencing is a specialized analysis that aims to identify and characterize genetic variations between an affected patient (proband) and a biological family member. The test includes DNA extraction, genome sequencing, and variant analysis for segregation. Segregation within family members will be noted in the proband's report.
• JAX Targeted Segregation Testing: Comparator targeted segregation testing is used to determine the presence/absence of variant(s) of interest in a patient's family members. The test includes DNA extraction and targeted Sanger sequencing or ddPCR. Results of segregation analysis will be noted in the proband's report.

• 1-2 mL of whole blood collected in 1 EDTA (lavender top) tube per patient.
  • For infants, a minimum of 0.5 mL is required.
• Samples must be submitted in the original EDTA collection tube with a minimum of two unique patient identifiers.
  • Germline Test Requisition

Samples that don't meet the above acceptance criteria may fail to generate useable data.

• Turnaround Time: ~30 days from sample receipt
• JAX Germline Sample Report

JAX Genome uses genomic DNA extracted from whole blood followed by a PCR-free library preparation using on-bead tagmentaton chemistry. Libraries are sequenced to a mean coverage of 30X using an Illumina NovaSeq 6000 instrument, which generates 151bp paired-end sequence reads. Sequencing reads are aligned to reference genome GRCh38 for variant calling. Variants are annotated and reviewed in the context of the patient's phenotype. Coding and noncoding regions of the genome are examined to detect SNVs, insertions/deletions, copy number changes and large structural variants.