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Possible Causes of Suboptimal Results


Remove WaveSense buffer from the refrigerator and bring to room temperature before beginning.
Vortex the beads and try not to create bubbles.
Place on CellCycler so they become well mixed prior to adding to the mixture.
It is recommended that bone marrow specimens be left at room temperature for stability of the sample. They may still be used for enrichment however this is not recommended.
Once the EpiSep HS caps have been removed the target area should be wet. It is critical to place them into the humidity chamber immediately prior to peeling back the caps. IF fixative evaporates quickly, the cell morphology is distorted.
Check to see if the slides have dried properly. The slides are to be dried in a humidity of 45-50%. Observe a slow drying process with a temperature of 25-27 degrees Celsius.
It is recommended that slides dry without manipulation. Do not place in front of a fan or use air from an aspirator. Cell morphology being preserved is critical since this is a direct prep method. Place slides standing up to air-dry.


Specimen Requirements and Issues


Not many labs have plasma cell percentages of the specimens prior to beginning enrichment. It is recommended to make a smear to obtain a morphological look at the plasma cell number. Use of the PlasmaCellQC Stain is highly specific for plasma cells. The smear also allows for examination of cell lies in the specimen.
WaveSense CD138+ enrichment can still be performed due to the sensitivity of the method. On average specimen volumes can be short. WaveSense CD138+ Targeted Cell enrichment sensitivity is 1%.
Recommendation One: Break up the clots manually with a pipette - DO NOT remove clots - This may be the only content that is available to get results.
Recommendation Two: After clots are manually broken up place the tube into a water bath at 37 degrees Celsius for 20 minutes to try and further dissolve the clot.
Since cells are happy in the RPMI and to save time, the RPMI may be substituted for the WaveSense Blocking Buffer.


General Protocol


NO.
The paramagnetic beads and Wave Sense Buffer are consistent regardless of the volume of the specimen.
On average you can successfully enrich between 5-7 days. Please follow our recommended reflex protocol that describes how to keep the specimen integrity. If no sample is left, it is possible to deprobe and already probed slide or save your supernatant from that patient, repeat the enrichment process.
NO.
The sensitivity of these beads is such that 3-5uls is standard as per the "Isolation of Target Cells from Bone Marrow" Tutorial. Larger amounts of beads can cause crowding in the target area of the EpiSep HS and mask cells of interest.
YES.
The supernatant may be used for additional cytogenetic or molecular downstream testing.
  • Place the cell suspension in 10 mL of Carnoy’s for storage.
  • When ready to make additional slides, centrifuge cell suspension 7 minutes at 1,000 RPM.
  • Remove the supernatant from the tube to just above the cell pellet.
  • Resuspend in fresh Carnoy’s fixative (3:1 methanol: acetic acid) according to the number of EpiSep slides you choose to prepare. NOTE: Make sure the volume is 1 mL per slide. If you want to make 3 slides, resuspend in 3mLs of Carnoy’s fixative (3:1 methanol: acetic acid).
We have seen examples where cells have been used as long as a month after fixation.


EpiSep Products


The white label is placed to the left. Slide the EpiSep HS into the MSD5 docks until you hear a “click”.
The EpiSep HS remains wet even though the well is dry. Remove the caps inside a humidified chamber by applying pressure to the white label side and peeling up.
The EpiSep HS remains wet even though the well is dry. Remove the supernatant from the loosely capped microvial on the opposite side of the magnet.
HELPFUL HINT: You may draw a line across the microvial to remind yourself to pipette away from the inner bead line.
The drain time of an EpiSep HS is 1 minute. The EpiSep HS can hold a total volume of 8 mLs. Apply pressure on the cap of the EpiSep HS as if you were to remove it but do not. Gently tap on the cap, which gets the flow moving and eliminates any possible air bubbles.
Click Here to view EpiSep trouble shooting tutorial.