A:
Blood collection for research usually follows clinical blood collection. Patients will not suffer with additional needle stick. Blood samples collected this way contain a low
number or none of the normal epithelial cells that could be introduced by needle sticking.
A:
There is no preference for the company brand of blood collection tubes. For example, BD Vacutainer™ tubes (Becton Dickinson, green top, each tube holds 6-mL) containing
Lithium or Sodium Heparin as anti-coagulants work well. Tubes using EDTA as anti-coagulants could reduce cell viability and are not recommended.
A:
Yes. Vita-Cap™ is designed to capture directly live rare cells from whole blood in one step. There are three types of rare cells emigrating from solid tissues in the blood;
i.e. cancer progenitor cells in patients’ blood, endothelial progenitor cells in all blood samples and fetal cells in maternal blood. These cells are rare and only reported
present in peripheral blood, ranging 5 to 500 cells per mL of blood.
A:
Yes. Approximately 1-3 mL of blood samples freshly collected from a blood collection tube should be transferred immediately into one Vita-Cap™ N6 tube (each tube added 2-mL
blood diluents containing anticoagulants) and subjected to cell enrichment by rotation at 10 rpm at 37°C for 3 hours for cancer cell attachment to occur. The rotation is best
done laying the tubes between 2 bars on a rotator. The 10 rpm is the speed of the circular churning motion of the tube around its long axis.
A:
No. Cancer or other rare cells rapidly lose their viability in blood collection tubes. Vita-Cap™ can also preserve the viability of its captured cells. Blood samples should
be processed immediately with Vita-Cap™ to prevent cell loss. After being captured by Vita-Cap™, cancer cells can remain viable longer. Vita-Cap™ with captured cancer cells
after removal of whole blood can be stored in tissue culture medium at 4 °C for a maximum of 24 hours before further processing and cell characterization.
A:
Protocols for use of our products that contain the specificity and sensitivity of Vita-Assay™ and Vita-Cap™ will be shipped with the products. The protocols are also available
as data sheets on the Research
Tools page on our website.
A:
The specificity and sensitivity of Vita-Assay™ and Vita-Cap™ have been determined using model experiments, in which 1 to 1,000 tumor cells are spiked into one mL of normal blood.
We also included a quality control of our products by directly comparing the capture efficiency of viable cancer cells from whole blood with current cell fractionation
methods (antibody-magnetic cell separation and density centrifugation). As available antibodies are directed against epithelial cells, we also use commonly used antibodies
against epithelial cells to detect cells recovered from our products. Our products generate highly enriched epithelial cell population that enables greater than 25 folds of
detection sensitivity of epithelial cells in blood than other methods.
A:
Isolated samples could be stored at -80 Celsius. Alternatively, cells can be first suspended by enzymes or DNA/RNA/Proteins extracted by appropriate buffers, and then stored
at -80 Celsius.
A:
CAM is able to enrich up to 1 million-fold rare cells derived from tissues (tumor, endothelium and bone marrow stem cells) in blood. For detection of CTCs, one can use CAM-enriched cells as primary sorting and identify CTCs by their positivity with epithelial lineage markers such as cytokeratins and epithelial cell adhesion molecules or by their negativity with CD45 (optional, as this has never occurred). For a sample containing 100 CTCs, there are approximately 1,000 CD45+ cells in a CAM-enriched portion that are negative for epithelial markers.
A:
Yes. In fact, CAM can enrich 10 times more tumor-like cells from bone marrow than blood from the same patient (judging by expression of epithelial lineage markers).
A:
There is a full manuscript published online by the journal
Gynocologic Oncology that describes some aspects of the technology and its
successful application in relating CTC count to patients' survival (PM:18954898).
You can find the manuscript Here. Another paper describing the functional phenotyping and genotyping of circulating tumor cells from patients with castration resistant prostate cancer has also been published (PM:19162393).
Q: From a hospital research governance and ethics point of view, could research samples be collected at the same time as patients' other routine bloods are being taken?
A: Blood collection for research usually follows clinical blood collection. Patients will not suffer with additional needle stick. Blood samples collected this way contain a low number or none of the normal epithelial cells that could be introduced by needle sticking.
Q: What type of blood collection tubes are recommended to take the blood prior to Vita-Cap™ processing?
A: There is no preference for the company brand of blood collection tubes. For example, BD VacutainerTM tubes (Becton Dickinson, green top, each tube holds 6-mL) containing Lithium or Sodium Heparin as anti-coagulants work well. Tubes using EDTA as anti-coagulants could reduce cell viability and are not recommended.
Q: Could the Vita-Cap™ isolate directly rare cancer cells from whole blood?
A: Yes. Vita-Cap™ is designed to capture directly live rare cells from whole blood in one step. There are three types of rare cells emigrating from solid tissues in the blood; i.e. cancer progenitor cells in patients’ blood, endothelial progenitor cells in all blood samples and fetal cells in maternal blood. These cells are rare and only reported present in peripheral blood, ranging 5 to 500 cells per mL of blood.
Q: Do blood samples need to be processed immediately?
A: Yes. Approximately 1-3 mL of blood samples freshly collected from a blood collection tube should be transferred immediately into one Vita-Cap™ N6 tube (each tube added 2-mL blood diluents containing anticoagulants) and subjected to cell enrichment by rotation at 10 rpm at 37°C for 3 hours for cancer cell attachment to occur. The rotation is best done laying the tubes between 2 bars on a rotator. The 10 rpm is the speed of the circular churning motion of the tube around its long axis.
Q: Could blood samples be stored in clinical blood collection tubes before being processed by Vita-Cap™?
A: No. Cancer or other rare cells rapidly lose their viability in blood collection tubes. Vita-Cap™ can also preserve the viability of its captured cells. Blood samples should be processed immediately with Vita-Cap™ to prevent cell loss. After being captured by Vita-Cap™, cancer cells can remain viable longer. Vita-Cap™ with captured cancer cells after removal of whole blood can be stored in tissue culture medium at 4 °C for a maximum of 24 hours before further processing and cell characterization.
Q: Do you have presentations or references relating your rare cell isolation assays?
A: Protocols for use of our products that contain the specificity and sensitivity of Vita-Assay™ and Vita-Cap™ will be shipped with the products. The protocols are also available as data sheets on the Research Tools <<link to http://www.vitatex.com/products_tools.asp>> page on our website.
Q: Have you compared the efficiency of your technology in the isolation of viable cancer cells with magnetic cell separation and Ficoll-Paque density centrifugation?
A: The specificity and sensitivity of Vita-Assay™ and Vita-Cap™ have been determined using model experiments, in which 1 to 1,000 tumor cells are spiked into one mL of normal blood. We also included a quality control of our products by directly comparing the capture efficiency of viable cancer cells from whole blood with current cell fractionation methods (antibody-magnetic cell separation and density centrifugation). As available antibodies are directed against epithelial cells, we also use commonly used antibodies against epithelial cells to detect cells recovered from our products. Our products generate highly enriched epithelial cell population that enables greater than 25 folds of detection sensitivity of epithelial cells in blood than other methods.
Q: Once the cells are washed can we store them at -80C without any freeze medium? Is it not necessary to use the enzyme if one is to isolate DNA/RNA/Proteins?
A: Isolated samples could be stored at -80 Celsius. Alternatively, cells can be first suspended by enzymes or DNA/RNA/Proteins extracted by appropriate buffers, and then stored at -80 Celsius.
Q: At which stage is it possible to freeze the cells: is it before or after enzymatic release of bound cells? Have you frozen the CTC tubes (with bound CTC on to the CAMs) to check the cell viability?
A: We recommend removing the cells (after enzymatic release) from the tube, and following the cell-freezing procedure
Q: What are the shipping conditions?
A: We ship this product at room temperature.
Q: Do you ship to foreign countries?
A: We have shipped this product to many foreign countries in Europe, Asia, as well as Australia using FedEx and DHL.
Q: How specific is CAM for viable tumor cells in blood?
A: CAM is able to enrich up to 1 million-fold rare cells derived from tissues (tumor, endothelium and bone marrow stem cells) in blood. For detection of CTCs, one can use CAM-enriched cells as primary sorting and identify CTCs by their positivity with epithelial lineage markers such as cytokeratins and epithelial cell adhesion molecules or by their negativity with CD45 (optional, as this has never occurred). For a sample containing 100 CTCs, there are approximately 1,000 CD45+ cells in a CAM-enriched portion that are negative for epithelial markers.
Q: Can we use CAM-initiated CTC sorting for the detection of "disseminated tumor cells" in bone marrow?
A: Yes. In fact, CAM can enrich 10 times more tumor-like cells from bone marrow than blood from the same patient (judging by expression of epithelial lineage markers).
Q: Are there peer-reviewed publications of CAM technology in CTC detection?
A: There is a full manuscript published online by the journal
Gynocologic Oncology that describes some aspects of the technology and its
successful application in relating CTC count to patients' survival (PM:18954898).
You can find the manuscript Here. Another paper describing the functional phenotyping and genotyping of circulating tumor cells from patients with castration resistant prostate cancer has also been published (PM:19162393).
