text.skipToContent text.skipToNavigation

AdnaTest LungCancer

For the enrichment and molecular characterization of circulating tumor cells (CTCs) from whole human blood.
  • Efficient isolation and detection of CTCs from whole blood
  • Molecular characterization of CTCs
  • High specificity and sensitivity
  • Minimize leukocyte bias with AdnaWash Leukocyte Reducer
AdnaTest LungCancer allows molecular characterization of CTCs by combining AdnaTest CTC-Select with AdnaTest LungCancerDetect using the Combination of Combinations Principle (COCP). AdnaTest CTC-Select is a highly specific immunomagnetic cell-selection system for enriching circulating tumor cells from peripheral blood. AdnaTest LungCancerDetect allows sensitive analysis of epithelial-mesenchymal transition (EMT) and lung cancer-associated gene expression in immunomagnetically enriched tumor cells by reverse transcription and PCR.
Cat No./ID: 395092
AdnaTest CTC-Select
¥96,500
Order Product
For immunomagnetic enrichment of circulating tumor cells (CTCs) from human whole blood for 12 preparations.
Cat No./ID: 396052
AdnaTest LungCancerDetect
¥83,000
Order Product
For 12 mRNA isolation from CTCs, reverse transcription and subsequent detection of epithelial-mesenchymal transition (EMT) lung cancer-associated gene expression using endpoint PCR.
Cat No./ID: 399911
AdnaMag-S
¥63,500
For 8 tubes, 1.5 ml
Cat No./ID: 399921
AdnaMag-L
¥63,500
For 8 tubes, 15 ml
0
Figure 1: Increased sensitivity in lung cancer.
In lung cancer, the AdnaTest CTC-Select provides antibodies against EpCam, Her2, EGFR, resulting in a strong increase of performance compared to detection based only on EpCam (Hanssen et al. (2016) Nature Sci Rep. 6, 28010).
1
Figure 2: Blood samples from healthy donors processed using PBS or AdnaWash.
2
Figure 3: A CTC captured by three antibodies coupled to magnetic beads.
3
Figure 4: AdnaTest CTC-Select: Immunomagnetic cell selection with multiple tumor associated antibodies.
In the first step, the CTCs in the blood are enriched (AdnaTest Select). This is achieved using antibody-coated magnetic particles (beads). Several antibodies are used, which bind with high specificity and affinity to the corresponding cancer cells. The enriched cells are lysed and subsequently purified several times to extract mRNA.
4
Figure 5: AdnaTest LungCancerDetect for analysis of various cancer-associated tumor markers.
This step consists of mRNA purification, reverse transcription and subsequent analysis of tumor associate markers by multiplex PCR. Resulting fragments are analyzed on a Bioanalyzer 2100.
5
Figure 6: AdnaTest LungCancerDetect results of duplex PCR samples using an Agilent 2100 Bioanalyzer.
The first lane shows the DNA size standard (DNA-Ladder). Samples 1 is positive for ALDH1 and GA733.2, sample 3 is positive for ALDH1 and sample 2 is negative. The PCR negative control  and the positive control (LungDetect) are shown in the last two lanes.
6
Figure 7: AdnaTest LungCancerDetect results of samples using an Agilent 2100 Bioanalyzer: EMT-2-associated gene expression.
The first lane shows the DNA size standard (DNA-Ladder). Sample 1 is negative. Sample 2 is positive for TWIST1, sample 3 is positive for Akt-2 and PI3Kα, and sample 4 is positive for PI3Kα and TWIST1. Signals for all three EMT-2 markers can be detected in sample 5 and actin is detected in samples 1 to 5. The PCR negative control (C-) and the positive control (EMT-2) are shown in the last two lanes.
Performance
AdnaTest LungCancer is a highly specific immunomagnetic cell selection system for enriching circulating tumor cells from peripheral blood which includes a highly sensitive RT-PCR for detecting lung cancer-associated mRNA markers. The specificity of the detection of AdnaTest LungCancer is 95%. Using tumor markers for lung cancer in spiking experiments, 30 tumor cells in 5 ml blood are detected at a recovery rate of 100% (Table below: AdnaTest LungCancer specificity and recovery).
AdnaTest LungCancer specificity and recovery
   ALDH1 GA733-2 All
 Non-spiked healthy donor samples  0/20 1/20 1/20
 Specificity  100% 95% 95%
 Spiked healthy donor samples  20/20 19/20 20/20
 Recovery 100% 95% 100%

In approximately 80% of all CTCs from lung cancer samples, EMT and tumor stem cell metabolism is observable. Analyzed CTCs were positive for at least one of the EMT markers: Twist1, Akt2 and PI3KCA. The AdnaTest LungCancer provides – in addition to anti-EpCam - antibodies against Her2 and EGFR, resulting in a strong increase of performance (see "Increased sensitivity in lung cancer"). To cover the aforementioned tumor markers, the AdnaTest LungCancer contains primer mixes for both the detection of EMT and lung cancer associated markers.

Findings with detection systems based solely on EpCam as capturing target reached only 15% positivity, whereas the AdnaTest CTC-Select uses anti-EpCam, anti-Her2 and anti-EGFR antibodies in combination. EpCam expression analysis enables differentiation of epithelial-like cells, EMT-like cells and cells with tumor stem cell characteristics.
Leukocyte reduction is critical to minimize leukocyte bias.

​The detection of tumor stemness-related markers like ALDH1, or EMT-related markers like Twist1, Akt2 or PI3KCα requires a significantly higher purity to eliminate a bias from contaminating leukocytes. The AdnaWash Leukocyte Reducer is a special buffer developed to reduce the leukocyte content during CTC enrichment. Using a CD45 endpoint PCR assay clearly demonstrates that the number of leukocytes is dramatically reduced using AdnaWash in the cell capturing procedure instead of PBS (see “Leukocyte reduction using AdnaWash”).
Principle
The AdnaTest’s successful CTC detection is based on the Combination of Combinations Principle (COCP). Each AdnaTest has a unique combination of tumor associated markers and an optimized combination of antibodies for cell selection. The combination of highly specific immunomagnetic cell selection system (using the optimized antibody combination, see “A CTC captured by three antibodies coupled to magnetic beads” ) and highly sensitive RT-PCR technology (using a combination of mRNA tumor markers) provides the highest degrees of analytical specificity and sensitivity (see "Increased sensitivity in lung cancer").
Procedure
The AdnaTest uses a two-step process (Select and Detect) to generate results within 5 hours (see “AdnaTest CTC-Select: Immunomagnetic cell selection with multiple tumor associated antibodies.” and “AdnaTest LungCancerDetect for analysis of various cancer-associated tumor markers.”). AdnaTest CTC-Select enables the immunomagnetic enrichment of tumor cells from whole blood via epithelial- and tumor-associated antigens. Antibodies against epithelial- and tumor-associated antigens are conjugated to magnetic beads for labeling of tumor cells. Labeled cells are extracted by a magnetic particle concentrator (AdnaMag-L and AdnaMag-S), and subsequently lysed. (see “AdnaTest Select: Immunomagnetic cell selection with multiple tumor-associated antibodies labeled to magnetic beads”). The cell lysate is then used in AdnaTest LungCancerDetect, step 2 of the procedure, to isolate mRNA.

AdnaTest LungCancerDetect contains Oligo (dT) 25-coated beads for the isolation of mRNA from the lysate of pre-enriched tumor cells. Reverse transcription results in cDNA that can subsequently be used as template for tumor cell detection and characterization by multiplex PCR (See "AdnaTest LungCancerDetect for analysis of various cancer-associated tumor markers.”).

​Multiplex PCR provides tumor-associated gene expression profiles for a variety of relevant tumor markers to ensure that the selected cells are cancer cells (see “AdnaTest LungCancerDetect results of samples using an Agilent 2100 Bioanalyzer”). The kit contains one primer mix to amplify cDNA target fragments of Twist, PI3Kα, Akt2 and a housekeeping gene as control (actin) (see "AdnaTest LungCancer specificity and recovery."). The PrimerMix LungDetect amplifies ALDH1 and EpCam (GA733-2) (below). The primers generate fragments of the following sizes:

Primer Mix EM-2 Detect
Target gene Fragment size
PI3Kα 551 bp
Akt-2 309 bp
TWIST1 201 bp
Actin 120 bp (internal PCR control)

Primer Mix LungDetect
Target Gene PCR fragment size
EpCam (GA733-2) 395 bp
ALDH1 161 bp

Importantly, the AdnaTest is an open platform that allows the profiling of additional custom selected mRNA targets for comprehensive biomarker discovery.
Applications
  • Isolation of CTCs from whole human blood
  • Molecular characterization of CTCs
  • Biomarker discovery
  • mRNA profiling
  • Liquid biopsy
  • Cancer research
fragment fix placeholder

Customers who bought these products also bought

  • Cat No./ID: 55284
    /jp/products//discovery-and-translational-research/dna-rna-purification/dna-purification/cell-free-dna/qiaamp-minelute-ccfdna-kits/

    QIAamp MinElute ccfDNA Midi Kit (50)

    For 50 preps (4 or 5 ml sample input volume each): QIAamp UCP MinElute Columns, QIAGEN Proteinase K, Magnetic Bead Suspension, Buffers, Bead Elution Tubes, and Collection Tubes (1.5 ml and 2 ml)
    ¥128,000