FAQ: Cell Based Assays

Q: Which CytoSelect™ Cell Migration Assay best fits my cell type?

A: Use the following table to determine which membrane pore size is best suited for your cell type:

Q: I have never run an assay using a Boyden Chamber. Would I have a better chance of success with your CytoSelect™ Cell Migration or Invasion kits if I use the 24-well or 96-well format?

A: If you are in the early stages of your migration or invasion studies and are still optimizing conditions, it is recommended that you start with the 24-well format. You can move to the 96-well format after conditions are optimized and you feel comfortable with the assay protocol.

Q: What is the difference between your product #CBA-101-COL where collagen is pre-coated on the underside (bottom) of the membrane, and your product #CBA 111-COL where collagen is coated on top of the membrane?

A: These products assay for different processes – cell migration and cell invasion, respectively. We offer four different types of cell migration assays, and you can get more information on the various definitions here. Our assay #CBA-101-COL is designed to measure haptotaxis, where immobilized ECM proteins act as the chemoattractant toward which cells migrate. Our assay #CBA-111-COL is designed to measure cell invasion, which is a more complex process that involves cell migration, degradation of the ECM, proteolysis, and movement of cells through the ECM to neighboring tissues. In this assay cells must be able to express active proteinases such as MMPs in order to digest and move through (i.e. invade) the coating on the top of the membrane.

Q: When using your CytoSelect™ Chemotaxis or Cell Invasion Assays, how can I guarantee that during the incubation with Cell Detachment Solution any non-migratory or non-invasive cells remaining in the top chamber will not pass through the membrane to the chamber below?

A:  The membrane pore size used in a Boyden chamber assay should always be smaller than the diameter of the cell to prevent passive movement of cells through the pores. The cell must rearrange its actin/myosin cytoskeleton structure in order to extend its leading edge and allow it to pass through the pores of the membrane. The 30 minute incubation with Cell Detachment Solution does not allow enough time for cells to do this.

Q: Do I need to use the entire Cell Migration or Invasion plate at once, or can I save unused wells for future tests?

A: Unused wells from our Radius™ Cell Migration Assays may be saved for future tests. The 24-well, 96-well, and 384-well plates may be used through a total of 3 migration experiment cycles. Unused wells of 24-well Boyden Chamber plates may be used in future experiments by removing unused cell culture inserts from the plate and storing them at 4ºC. 96-well Boyden Chamber Migration plates are designed to be used in their entirety, although very careful protection of unused wells may allow them to be used in future experiments. Migration plates should be sterilized by exposure to ultraviolet light for 20 minutes prior to any subsequent use. 96-well Cell Invasion plates should be used in their entirety at once, because repeated incubations could compromise the integrity of the membrane coating.
 

Q: You say manual counting of cells is eliminated when using the CytoSelect™ Cell Transformation Assay (Cell Recovery Compatible), but how can I quantify my cells and still recover them for further analysis?

A: All of our CytoSelect™ Cell Transformation Assays include a quantitation step that eliminates manual cell counting. However, this quantitation does require cell lysis. When using our Cell Recovery Compatible Assay (#CBA-135 or CBA-140), it is recommended that duplicate wells (or more) be run for each sample. This will allow you to quantify cells in one well and then recover the cells from the duplicate well.

Q: I’m interested in your CytoSelect™ 96-Well Cell Transformation Assay (Soft Agar Colony Formation). Can the CyQuant dye in this assay distinguish living cells from dead cells in agarose?

A: The CyQuant dye counts cells by measuring DNA/RNA; it cannot distinguish living cells from dead cells. However, our CytoSelect™ Cell Transformation Assay, Cell Recovery Compatible #CBA-135 uses MTT Solution to count cells and allows viable transformed cells to be easily recovered for further culturing and testing. If you run duplicate samples in this assay, you could use one well for MTT cell counting and the other for cell recovery which would allow you to do further analysis of your viable cells.

Q: I’m interested in using your CytoSelect™ 96-Well Cell Transformation Assay. I noticed that your kit comes with DMEM Solution, is it possible to use my own cell culture media instead of the DMEM provided?

A: Yes; you can use your own media instead of the DMEM we provide in the kit, but only if you are able to make your complete medium at a 2X concentration. You will mix your cells/media 1:1 with the agar matrix which will leave you with a final concentration of 1X. If you are unable to obtain or make your own media at a 2X concentration, then we suggest using the DMEM we provide.

Q: Do your CytoSelect™ Microfluidic Biochips require a specific kind of syringe pump?

A: Syringe pumps that are compatible include those from Harvard Apparatus, World Precision Instruments, and KD Scientific, but would likely work with other brands also. The main consideration when choosing a syringe pump is to ensure it has the capability to both “push” and “pull”, i.e. dispense and aspirate. Some pumps will only dispense.

Q: Can I buy just the inserts from the CytoSelect™ 24-Well Wound Healing Assay and use them with my own plates?

A: The inserts used in our CytoSelect™ Wound Healing Assay are specially molded to create a precise, consistent gap from well to well. The inserts are designed to be perfectly flat at the bottom. We have screened many commercially available 24-well plates and found that most of them have wells that are not sufficiently flat to create good contact with the bottom of the insert. To ensure quality results with this assay, we only guarantee results when using our inserts with the plates included in our kit. Inserts are not available for separate purchase.

Q: Can the inserts from your CytoSelect™ 24-Well Wound Healing Assay be autoclaved and re-used?

A: No, the inserts are not autoclavable and will be significantly distorted if heated.

Q:  I am interested in using the CytoSelect™ 96-Well Phagocytosis Assay with Zymosan Substrate. Could you recommend an opsonizing reagent for Zymosan particles?

A: Zymosan is from yeast, which is a natural pathogen for phagocytes.  You don't need to opsonize the particles before use in this assay, but if you prefer to do so the best method is serum. For example, if you are studying human cell phagocytosis, we recommend using human serum for opsonization.

Q:  What types of cells may be used in your Collagen-based Cell Contraction Assay?

A: Any cell type may be used as long as the cells can be cultured in 3D collagen.  Besides fibroblasts, the collagen model may be used to study endothelial cells, epithelial cells, muscle cells and heart cell contraction.

Q: How should I report my results obtained from using your Endothelial Tube Formation Assay (In Vitro Angiogenesis)?

A: There are two ways to report your results: by tube length or by number of branch points.