Meet the Expert on the ClearLLab 10C System
Sandra Hernandez answers your questions on the ClearLLab 10C system, the only integrated leukemia and non-Hodgkin’s lymphoma immunophenotyping solution for lymphoid and myeloid lineages.
Sandra Hernandez
Sandra is a manager in the global clinical flow cytometry marketing team and is responsible for commercial marketing activities for on-market clinical products. She joined Beckman Coulter over five years ago. Sandra is a licensed Medical Technologist Supervisor and holds a qualification in Flow Cytometry from ASCP. Prior to joining Beckman Coulter, Sandra was the supervisor of the Clinical Flow Cytometry and Molecular Pathology laboratory at the University of Miami in Miami, Florida.
With reference to residual cells as an internal reference, is there a possibility that a sample has no normal residual cells?
Yes, that is a possibility. When I was at the University of Miami, you would look at the sample you ran before, which would possibly have these residual cells -- or possibly a sample that you would run after -- because maybe you come across a leukemia that has 90% blast, which means there are no normal samples or very few normal samples. As you use these panels that are validated, you can justify that control by looking at the staining of samples you ran before or after. That gives you the peace of mind of knowing that your panels are working as they should.
With ClearLLab 10C, should you use only IOTest 3 lysing and fixative solutions, or can you use other lysing reagents such as VersaLyse?
For the moment, ClearLLab 10C reagents have been validated for use with IOTest 3 lysing reagents. Our R&D Team conducted studies to compare Versalyse with the IOTest 3, and there was much better separation using the IOTest 3 reagents, which are also not as harsh on cells because they’re based on ammonium chloride. Ammonium chloride is a gentle lyse that helps preserve the fragile blasts and the extremely fragile plasma cells. What we would like to do is align with the ClearLLab LS (available only in Europe) and add the IOTest 3 lyse to that, which would provide one method across all the ClearLLab processes.
How do you download the Levy-Jennings templates used with ClearLLab 10C?
These can be downloaded from Beckman.com. These are lot-specific, so for each new lot, new templates should be downloaded. The analysis templates are also there, plus the system guide, which walks you through that download process and an addendum with instructions to find the templates for the controls. Everything you need to run the controls and run the analysis is available online.
Are the ClearLLab panels sufficient for identifying MPNs that don't show blast populations?
As ClearLLab has many different markers, we can look at the changes and different patterns of expression to identify some of the myeloproliferative neoplasms (MPNs) even if they don't have blasts. There are several publications that use that type of workflow to enable identification of MPNs. There isn’t a specific marker for an MPN, but you can look at some of the changes in the patterns of expression that can lead you to a differential diagnosis.
Can we use these IQC materials for non hemato-oncology assays e.g., lymphocyte subsets, ALPS panels etc.?
ClearLLab 10C is an IVD system and should be used only within its intended use. If it's within the ClearLLab 10C System claims, then it can be used. Deviating from IVD intended use, however, means you are working outside of these claims, and you must evaluate and validate it, as the IVD status no longer applies.
I'm using a DxFLEX five-color with a blue laser; should I use your immune-control cells in my machine?
No. The control cells are validated for use on Navios and Navios EX flow cytometers only.
Are ClearLLab 10C and ClearLLab controls suitable for Minimal Residual Disease (MRD) research?
ClearLLab 10C reagents and controls are IVD and used as an aid to diagnosis. They have not been validated for MRD in the clinical setting.
What is the form of ClearLLab controls?
ClearLLab controls are whole blood process controls that are processed in parallel, using the same procedure used for processing the sample. It's a true process control as opposed to lyophilized, which doesn't have the same matrix seen with whole blood controls.
What is the shelf-life of ClearLLab reagents?
ClearLLab has a 12-month shelf life, and DURAClone RUO reagents have up to 18 months of shelf life. This provides stability and is especially useful if you want to maintain the same lot. Reagents with long shelf lives require less work in the lab, as users no longer have to do side-by-side comparisons of old versus new lots, thereby improving overall workflow efficiency.
Is ClearLLab 10C authorized for use in North America and Europe?
ClearLLab 10C is the only FDA authorized IVD panel and control for leukemia and lymphoma analysis. It is also CE-marked for Europe. ClearLLab will be one of the first assays to go through the new IVDR regulatory process.
How can we use the ClearLLab 10C panel to identify Hematogone or B-cell / ALL?
In the B cell tube, you have CD34, CD19, CD10 and CD38, and with these markers you can definitely see if you have hematogones and the precursor B cells. You can also differentiate those into immature and mature B cell markers, because they lose some markers and gain other B cell markers in their maturity.
Are there any additional training resources for ClearLLab?
The ClearLLab casebook is an excellent training tool. It introduces new ClearLLab users to different populations of normal and abnormal samples.
Flow cytometry is about pattern recognition, and whenever you introduce a new panel, you need to learn to recognize the patterns seen with different antibody combinations. This training resource is an excellent tool for anybody who’s new to the lab or is not familiar with analysis of ClearLLab reagents.
The book contains more than 25 different cases. It starts with illustrations of normals from each different specimen type: bone marrow, peripheral blood and lymph node, to help readers understand what normal looks like.
Only after we know what normal looks like can we distinguish the abnormals. The Casebook then divides abnormals into the different lineage-specific types, looking at immature and mature examples within the B-cell, T-cell and Myeloid types.
The interpretations covered in the ClearLLab casebook were conducted by Professor Brent Wood and his team at the University of Washington. There’s an introduction to each case, with clinical information and demographics which helps demonstrate how cases typically present themselves. Each histogram has extensive annotations, which describe the normal and abnormal. Each case concludes with any additional clinical information required for interpretation. Technologists and supervisors alike can use the casebook as a reference for scenarios they might encounter when running the ClearLLab panel.
What about TDT and HLA-DR markers? Are they included with CD117 and CD34?
We have HLA-DR in both Myeloid tubes M1 and M2. TDT is not part of the panel and should be run as a reflex to confirm lineage if not apparent from the panel’s marker staining. The M2 tube has CD34/CD117 and HLA-DR.
What is the difference between ClearLLab M1 and M2 tubes, and which one should be used first?
We recommend using both tubes, as they cover granulocyte and monocyte markers. Also, there is no marker in M1 or M2 tubes for M6 or M7 subtypes. As these are low-frequency Myeloid leukemias, the specific markers for erythroid precursors and platelet antibodies were not included in the ClearLLab Myeloid tubes.
For T-cell tubes, you add only CD56 for NKs; does adding CD16 help here (e.g. to detect NK sub-populations)?
We have CD16 in the M1 tube, which can be helpful. If an NK neoplasm is suspected, then a more specific NK reflex tube can be done.
How often should labs perform compensation or calibration on these systems?
Under three specific circumstances: (1) on initial set up, (2) when daily QC fails, and (3) when using a new lot of Flow-Set beads ClearLLab 10C panels include compensation reagents, a condensation kit and compensation beads.
Is TDT included in the panel?
No. If lineage remains unclear, TDT should be set up as a reflex test.
How often are MPO, CD22, CD11c and CD1a required to establish a final diagnosis in acute leukemias?
This will depend on your lab's workflow, patient population, and the frequency of acute leukemias in that population.
Because these tubes do not contain cytoplasmic staining tubes, how do we establish lineage for T-lymphoblasts, given that cytoplasmic CD3 is the lineage marker for them?
This would be set up as a confirmation if CD3 population is negative for surface expression.
ClearLLab LS
Because TDT is not in the tube, is it possible to miss CD34-negative, TDT-positive leukemias by the LST?
You should be able to identify a dim CD45 population by side scatter and then reflex the TDT to establish lineage, if required.
Can we use ClearLLab 10C reagents with cerebrospinal fluid?
No. The ClearLLab LS tube has been validated only for use with bone marrow, peripheral blood and lymph node specimens.
Can the screening tube be used to identify plasma cell disorders?
No. Plasma cells are not part of the ClearLLab LS screening tube.
