Ms and Bone Marrow Transplantation, University Hospital in Wroclaw (Lab2); Department
Ms and Bone Marrow Transplantation, University Hospital in Wroclaw (Lab2); Department of Experimental Hematooncology, Health-related University of Lublin (Lab3) and as a Coordinating Laboratory, Laboratory of Immunophenotyping, Institute of Hematology and Transfusion Medicine in Warsaw (Lab4). In 2019, an electronic survey was performed, aimed at verifying compliance on the MRD assays protocols of your MM MRD assay in every laboratory. The participants had been requested to supply categorized information concerning the MFC MRD assessment process such as the type of instrument applied, flow cytometer settings, antibody panels, staining procedure conditions, too as the experience in the staff in performing MRD tests in MM. The outcomes in the survey had been analyzed by the Coordinating Laboratory. Considering that all laboratories confirmed the usage of the EuroFlow-adapted sample preparation protocol, inside the initial phase of our study, we decided to standardize instrument settings in line with EuroFlow procedures. The required reagents and antibodies have been acquired and distributed for the participants by the Coordinating Laboratory. The second phase of the study aimed at assessing the inter-laboratory variability of myeloma Pc measurements inside the same BM samples, evaluated as outlined by local protocols for MRD assessment in MM. In 2020, 12 BM Compound 48/80 Purity & Documentation samples (S1 12), had been prepared and distributed by the Coordinating Laboratory to the participating laboratories in three rounds. Immediately after evaluating the samples, the sites provided flow cytometry data files (fcs.) to the Coordinating Laboratory for evaluation. Central evaluation aimed also at figuring out the intra-assay variation (repeatability) and inter-laboratory comparison of your fluorescence intensity of your labeled antigens on regular plasma cells (PCs) obtained soon after instrument standardization. The third phase with the study aimed at evaluating the inter-operator variability in MRD determination and MM plasma cell immunophenotype classification in the identical cytometric information files. Raw cytometric information files (fcs.) of 13 sufferers with distinctive MRD status (SA1 A13) have been electronically distributed to the participant laboratories by the Coordinating Laboratory. Soon after every study phase, the results with the comparisons have been communicated to the participant laboratories and discussed. two.two. Instruments Setup Standardization Standardization of all flow cytometers settings was Decanoyl-L-carnitine Purity & Documentation performed by implementation of the EuroFlow Standard Operating Protocol (SOP) for instrument setup and compensation for FASCCanto II and FACSLyric, respectively (www.euroflow.org, accessed on 7 October 2021) [25]. So as to setup photomultiplier (PMT) voltages in FACSCantoII instruments, we made use of median fluorescence intensity (MdFI) in the 7th reference peak of Rainbow beads calibration particles (Spherotech Inc., Lake Forest, IL, USA), EuroFlow-validated lot number EAK01. To setup standardized and comparable fluorescence measurements in FACSLyric flow cytometers, EuroFlow has defined specific tube target values (TTV) for every emission filter and fluorochrome. The acceptable tube settings and/or assays for FASCLyric are accessible around the EuroFlow site (www.euroflow.org, accessed on 7 October 2021). Just before acquisition from the study samples, Rainbow beads with the similar lot quantity were acquired, so that you can monitor each instrument functionality between study rounds. Furthermore,Diagnostics 2021, 11,4 ofparticipants had been asked to obtain and record Rainbow beads on their routinely.
