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Genetic profiling of tumours using both circulating free DNA and circulating tumour cells isolated from the same preserved whole blood sample.

Rothwell DG, Smith N, Morris D, Leong HS, Li Y, Hollebecque A, Ayub M, Carter L, Antonello J, Franklin L, Miller C, Blackhall F, Dive C, Brady G - Mol Oncol (2015)

Bottom Line: However, accurate assessment of both CTCs and ctDNA requires all blood cells to be maintained intact until samples are processed.We demonstrate that yields of circulating free DNA (cfDNA) obtained from whole blood CellSave samples are equivalent to those obtained from conventional EDTA plasma processed within 4 h of blood draw.Targeted and genome-wide NGS revealed comparable DNA quality and resultant sequence information from cfDNA within CellSave and EDTA samples.

View Article: PubMed Central - PubMed

Affiliation: Nucleic Acid Biomarker Laboratory, Clinical Experimental Pharmacology Group, CR-UK Manchester Institute, University of Manchester, M20 4BX, UK. Electronic address: dominic.rothwell@cruk.mancehster.ac.uk.

No MeSH data available.


Related in: MedlinePlus

A. Yields of cfDNA from duplicate clinical samples collected in EDTA and CellSave bloods from a cohort of 11 SCLC and 34 melanoma patients. No significant difference was found between each collection type in both cohorts. B. Mutations identified in five SCLC patient samples using a targeted NGS approach. Germline gDNA, EDTA cfDNA and CellSave cfDNA was analysed for each patient. Mutations were called with read counts >200 and frequency >10%. Mutated samples are indicated by red fill with WT alleles indicated by green fill.
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fig3: A. Yields of cfDNA from duplicate clinical samples collected in EDTA and CellSave bloods from a cohort of 11 SCLC and 34 melanoma patients. No significant difference was found between each collection type in both cohorts. B. Mutations identified in five SCLC patient samples using a targeted NGS approach. Germline gDNA, EDTA cfDNA and CellSave cfDNA was analysed for each patient. Mutations were called with read counts >200 and frequency >10%. Mutated samples are indicated by red fill with WT alleles indicated by green fill.

Mentions: CellSave vacutainers are routinely used for CTC enumeration using the CellSearch® platform and molecular analysis of CTCs retrieved from CellSearch® cartridges can be achieved using both focused and genome wide NGS (Hodgkinson et al., 2014, Gasch et al., 2013, Heitzer et al., 2013). Since the CellSearch® system requires 7.5 ml blood input and the CellSave vacutainer can hold up to 10 ml there is often surplus blood, which can be used for additional analyses. To test the suitability of CellSave for cfDNA analysis of clinical samples, we compared yields of cfDNA obtained from surplus CellSave blood to yields of cfDNA obtained from sample obtained from a parallel EDTA blood sample processed to plasma within 4 h from two clinical cohorts. Analysis of 11 SCLC and 34 melanoma patient samples showed comparable yields of patient cfDNA from 4 h EDTA plasma (hereafter referred to as standard EDTA) to cfDNA isolated from CellSave blood kept at room temperature for up to 96 h (Figure 3A). This mirrored the results from the HNV experiment and showed CellSave blood to be stable source of both cfDNA and CTCs for clinical sample analysis.


Genetic profiling of tumours using both circulating free DNA and circulating tumour cells isolated from the same preserved whole blood sample.

Rothwell DG, Smith N, Morris D, Leong HS, Li Y, Hollebecque A, Ayub M, Carter L, Antonello J, Franklin L, Miller C, Blackhall F, Dive C, Brady G - Mol Oncol (2015)

A. Yields of cfDNA from duplicate clinical samples collected in EDTA and CellSave bloods from a cohort of 11 SCLC and 34 melanoma patients. No significant difference was found between each collection type in both cohorts. B. Mutations identified in five SCLC patient samples using a targeted NGS approach. Germline gDNA, EDTA cfDNA and CellSave cfDNA was analysed for each patient. Mutations were called with read counts >200 and frequency >10%. Mutated samples are indicated by red fill with WT alleles indicated by green fill.
© Copyright Policy - CC BY-NC-ND
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4834815&req=5

fig3: A. Yields of cfDNA from duplicate clinical samples collected in EDTA and CellSave bloods from a cohort of 11 SCLC and 34 melanoma patients. No significant difference was found between each collection type in both cohorts. B. Mutations identified in five SCLC patient samples using a targeted NGS approach. Germline gDNA, EDTA cfDNA and CellSave cfDNA was analysed for each patient. Mutations were called with read counts >200 and frequency >10%. Mutated samples are indicated by red fill with WT alleles indicated by green fill.
Mentions: CellSave vacutainers are routinely used for CTC enumeration using the CellSearch® platform and molecular analysis of CTCs retrieved from CellSearch® cartridges can be achieved using both focused and genome wide NGS (Hodgkinson et al., 2014, Gasch et al., 2013, Heitzer et al., 2013). Since the CellSearch® system requires 7.5 ml blood input and the CellSave vacutainer can hold up to 10 ml there is often surplus blood, which can be used for additional analyses. To test the suitability of CellSave for cfDNA analysis of clinical samples, we compared yields of cfDNA obtained from surplus CellSave blood to yields of cfDNA obtained from sample obtained from a parallel EDTA blood sample processed to plasma within 4 h from two clinical cohorts. Analysis of 11 SCLC and 34 melanoma patient samples showed comparable yields of patient cfDNA from 4 h EDTA plasma (hereafter referred to as standard EDTA) to cfDNA isolated from CellSave blood kept at room temperature for up to 96 h (Figure 3A). This mirrored the results from the HNV experiment and showed CellSave blood to be stable source of both cfDNA and CTCs for clinical sample analysis.

Bottom Line: However, accurate assessment of both CTCs and ctDNA requires all blood cells to be maintained intact until samples are processed.We demonstrate that yields of circulating free DNA (cfDNA) obtained from whole blood CellSave samples are equivalent to those obtained from conventional EDTA plasma processed within 4 h of blood draw.Targeted and genome-wide NGS revealed comparable DNA quality and resultant sequence information from cfDNA within CellSave and EDTA samples.

View Article: PubMed Central - PubMed

Affiliation: Nucleic Acid Biomarker Laboratory, Clinical Experimental Pharmacology Group, CR-UK Manchester Institute, University of Manchester, M20 4BX, UK. Electronic address: dominic.rothwell@cruk.mancehster.ac.uk.

No MeSH data available.


Related in: MedlinePlus