Cytometry Part B: Clinical Cytometry最新文献

{"title":"Flow cytometry of DNMT1 as a biomarker of hypomethylating therapies","authors":"Philip G. Woost,&nbsp;Basem M. William,&nbsp;Brenda W. Cooper,&nbsp;Masumi Ueda Oshima,&nbsp;Folashade Otegbeye,&nbsp;Marcos J. De Lima,&nbsp;David Wald,&nbsp;Reda Z. Mahfouz,&nbsp;Yogen Saunthararajah,&nbsp;Tammy Stefan,&nbsp;James W. Jacobberger","doi":"10.1002/cyto.b.22158","DOIUrl":"10.1002/cyto.b.22158","url":null,"abstract":"<p>The 5-azacytidine (AZA) and decitabine (DEC) are noncytotoxic, differentiation-inducing therapies approved for treatment of myelodysplastic syndrome, acute myeloid leukemias (AML), and under evaluation as maintenance therapy for AML postallogeneic hematopoietic stem cell transplant and to treat hemoglobinapathies. Malignant cell cytoreduction is thought to occur by S-phase specific depletion of the key epigenetic regulator, DNA methyltransferase 1 (DNMT1) that, in the case of cancers, thereby releases terminal-differentiation programs. DNMT1-targeting can also elevate expression of immune function genes (HLA-DR, MICA, MICB) to stimulate graft versus leukemia effects. In vivo, there is a large inter-individual variability in DEC and 5-AZA activity because of pharmacogenetic factors, and an assay to quantify the molecular pharmacodynamic effect of DNMT1-depletion is a logical step toward individualized or personalized therapy. We developed and analytically validated a flow cytometric assay for DNMT1 epitope levels in blood and bone marrow cell subpopulations defined by immunophenotype and cell cycle state. Wild type (WT) and DNMT1 knock out (DKO) HC116 cells were used to select and optimize a highly specific DNMT1 monoclonal antibody. Methodologic validation of the assay consisted of cytometry and matching immunoblots of HC116-WT and -DKO cells and peripheral blood mononuclear cells; flow cytometry of H116-WT treated with DEC, and patient samples before and after treatment with 5-AZA. Analysis of patient samples demonstrated assay reproducibility, variation in patient DNMT1 levels prior to treatment, and DNMT1 depletion posttherapy. A flow-cytometry assay has been developed that in the research setting of clinical trials can inform studies of DEC or 5-AZA treatment to achieve targeted molecular pharmacodynamic effects and better understand treatment-resistance/failure.</p>","PeriodicalId":10883,"journal":{"name":"Cytometry Part B: Clinical Cytometry","volume":"106 1","pages":"11-24"},"PeriodicalIF":3.4,"publicationDate":"2024-02-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/cyto.b.22158","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139722011","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Enhancing HLA-B27 antigen detection: Leveraging machine learning algorithms for flow cytometric analysis.","authors":"Sándor Baráth, Parvind Singh, Zsuzsanna Hevessy, Anikó Ujfalusi, Zoltán Mezei, Mária Balogh, Marianna Száraz Széles, János Kappelmayer","doi":"10.1002/cyto.b.22164","DOIUrl":"https://doi.org/10.1002/cyto.b.22164","url":null,"abstract":"<p><p>As the association of human leukocyte antigen B27 (HLA-B27) with spondylarthropathies is widely known, HLA-B27 antigen expression is frequently identified using flow cytometric or other techniques. Because of the possibility of cross-reaction with off target antigens, such as HLA-B7, each flow cytometric technique applies a \"gray zone\" reserved for equivocal findings. Our aim was to use machine learning (ML) methods to classify such equivocal data as positive or negative. Equivocal samples (n = 99) were selected from samples submitted to our institution for clinical evaluation by HLA-B27 antigen testing. Samples were analyzed by flow cytometry and polymerase chain reaction. Features of histograms generated by flow cytometry were used to train and validate ML methods for classification as logistic regression (LR), decision tree (DT), random forest (RF) and light gradient boost method (GBM). All evaluated ML algorithms performed well, with high accuracy, sensitivity, specificity, as well as negative and positive predictive values. Although, gradient boost approaches are proposed as high performance methods; nevertheless, their effectiveness may be lower for smaller sample sizes. On our relatively smaller sample set, the random forest algorithm performed best (AUC: 0.92), but there was no statistically significant difference between the ML algorithms used. AUC values for light GBM, DT, and LR were 0.88, 0.89, 0.89, respectively. Implementing these algorithms into the process of HLA-B27 testing can reduce the number of uncertain, false negative or false positive cases, especially in laboratories where no genetic testing is available.</p>","PeriodicalId":10883,"journal":{"name":"Cytometry Part B: Clinical Cytometry","volume":" ","pages":""},"PeriodicalIF":3.4,"publicationDate":"2024-02-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139722010","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Role of flow cytometric immunophenotyping in the diagnosis of breast implant-associated anaplastic large cell lymphoma: A 6-year, single-institution experience","authors":"Alexander Chan,&nbsp;Romany Auclair,&nbsp;Qi Gao,&nbsp;Paola Ghione,&nbsp;Steven Horwitz,&nbsp;Ahmet Dogan,&nbsp;Mikhail Roshal,&nbsp;Oscar Lin","doi":"10.1002/cyto.b.22162","DOIUrl":"10.1002/cyto.b.22162","url":null,"abstract":"<p>Breast implant-associated anaplastic large cell lymphoma (BIA-ALCL) is an uncommon mature T-cell neoplasm occurring in patients with textured breast implants, typically after 7–10 years of exposure. Although cytopathologic or histopathologic assessment is considered the gold standard diagnostic method for BIA-ALCL, flow cytometry (FC)-based immunophenotyping is recommended as an adjunct test. However, the diagnostic efficacy of FC is not well reported. We reviewed 290 FC tests from breast implant pericapsular fluid and capsule tissue from 182 patients, including 16 patients with BIA-ALCL over a 6-year period, calculating diagnostic rates and test efficacy. FC showed an overall sensitivity of 75.9%, specificity of 100%, and negative and positive predictive values of 95.4% and 100%, respectively. Blinded expert review of false-negative cases identified diagnostic pitfalls, improving sensitivity to 96.6%. Fluid samples had better rates of adequate samples for FC testing compared with tissue samples. Paired with FC testing of operating room (OR)-acquired fluid samples, capsulectomy FC specimens added no diagnostic value in patients with concurrent fluid samples; no cases had positive capsule FC with negative fluid FC. Fluid samples are adequate for FC testing more often than tissue. Capsule tissue FC specimens do not improve FC efficacy when paired with OR-acquired fluid FC samples and are often inadequate samples. FC is 100% specific for BIA-ALCL and can serve as a confirmatory test but should not be the sole diagnostic method. Awareness of sample-specific diagnostic pitfalls greatly improves the sensitivity of BIA-ALCL testing by FC.</p>","PeriodicalId":10883,"journal":{"name":"Cytometry Part B: Clinical Cytometry","volume":"106 2","pages":"117-125"},"PeriodicalIF":3.4,"publicationDate":"2024-01-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139650467","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"ROR1 expression in mature B lymphoid neoplasms by flow cytometry","authors":"Flávia Arandas de Sousa,&nbsp;Nádila Magalhães Millan,&nbsp;Rodolfo Patussi Correia,&nbsp;Andressa da Costa Vaz,&nbsp;Daniela Schimidell,&nbsp;Priscila Carmona Miyamoto,&nbsp;Marilia Sandoval Passaro,&nbsp;Bruna Garcia Nogueira,&nbsp;Elizabeth Xisto Souto,&nbsp;Nydia Strachman Bacal,&nbsp;Laiz Camerão Bento","doi":"10.1002/cyto.b.22157","DOIUrl":"10.1002/cyto.b.22157","url":null,"abstract":"<p>Immunophenotyping by flow cytometry is an integral part of the diagnosis and classification of leukemias/lymphomas. The expression of ROR1 associated with chronic B lymphocytic leukemia (CLL) is well described in the literature, both in its diagnosis and in the follow-up of minimal residual disease (MRD) research, however, there are few studies regarding the expression pattern of ROR1 in other subtypes of mature B lymphoid neoplasms. With the aim of evaluating the expression of ROR1 and associating it with the expression of other important markers for the differentiation of mature B lymphoid neoplasms (MBLN), 767 samples of cases that entered our laboratory for immunophenotyping with clinical suspicion of MBLN were studied. ROR1 expression is predominant in CD5+/CD10− neoplasms. Overall, positive ROR1 expression was observed in 461 (60.1%) cases. The CD5+/CD10− group had a significantly higher proportion of ROR1 positive samples (89.9%) and more brightly expressed ROR1 than the other groups. Our results highlight the importance of evaluating ROR1 expression in the diagnosis of MBLN to contribute to the differential diagnosis, and possibly therapy of mainly CLL, and indicate that this marker could be considered as a useful addition to immunophenotypic panels, particularly for more challenging cases.</p>","PeriodicalId":10883,"journal":{"name":"Cytometry Part B: Clinical Cytometry","volume":"106 1","pages":"74-81"},"PeriodicalIF":3.4,"publicationDate":"2024-01-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139564045","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Maturational dyssynchrony in benign B-cell precursors following lymphocyte depleting chemotherapy: A potential pitfall for B-lymphoblastic leukemia minimal/measurable residual disease (MRD) flow cytometry analysis","authors":"Alexander Placek,&nbsp;Brian Lockhart,&nbsp;Karin P. Miller,&nbsp;Gerald B. Wertheim,&nbsp;Shannon L. Maude,&nbsp;Brent L. Wood,&nbsp;Alexandra E. Kovach","doi":"10.1002/cyto.b.22161","DOIUrl":"10.1002/cyto.b.22161","url":null,"abstract":"","PeriodicalId":10883,"journal":{"name":"Cytometry Part B: Clinical Cytometry","volume":"106 2","pages":"138-141"},"PeriodicalIF":3.4,"publicationDate":"2024-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139511522","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Deciphering stage 0 hematogones by flow cytometry in follow-up bone marrow samples of pediatric B—Acute lymphoblastic leukemia cases: A potential mimicker of residual disease after anti CD19 therapy","authors":"Thulasi Raman Ramalingam,&nbsp;Lakshman Vaidhyanathan,&nbsp;Anurekha Muthu,&nbsp;Venkateswaran Vellaichamy Swaminathan,&nbsp;Ramya Uppuluri,&nbsp;Revathi Raj","doi":"10.1002/cyto.b.22159","DOIUrl":"10.1002/cyto.b.22159","url":null,"abstract":"<p>CD19 is frequently targeted for immunotherapy in B cell malignancies, which may result in loss of CD19 expression in leukemic cells as an escape mechanism. Stage 0 hematogones (Hgs) are normal CD19-negative very early B cell precursors that can be potentially mistaken for CD19 negative residual leukemic cells by flow cytometry (FCM) in B cell acute lymphoblastic leukemia (BCP-ALL) cases treated with anti CD19 therapy. Our main objective was to characterize and study the incidence of stage 0 hematogones in follow-up bone marrow samples of pediatric BCP-ALL cases. We analyzed the flow cytometry standard files of 61 pediatric BCP-ALL cases treated with conventional chemotherapy and targeted anti-CD19 therapy, for identifying the residual disease and normal B cell precursors including stage 0 Hgs. A non-CD19 alternate gating strategy was used to isolate the B cells for detecting the residual disease and stage 0 Hgs. The stage 0 Hgs were seen in 95% of marrow samples containing CD19+ Hgs. When compared with controls and posttransplant marrow samples, the fraction of stage 0 Hgs was higher in patients receiving anti CD19 therapy (<i>p</i> = 0.0048), but it was not significant when compared with patients receiving chemotherapy (<i>p</i> = 0.1788). Isolated stage 0 Hgs are found in samples treated with anti-CD19 therapy simulating CD19 negative residual illness. Our findings aid in understanding the stage 0 Hgs and its association with CD19+ Hgs in anti CD19 therapy and conventional chemotherapy. This is crucial as it can be potentially mistaken for residual disease in patients treated with anti CD19 therapy.</p>","PeriodicalId":10883,"journal":{"name":"Cytometry Part B: Clinical Cytometry","volume":"106 2","pages":"92-98"},"PeriodicalIF":3.4,"publicationDate":"2024-01-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139502365","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Standardization of flow cytometric detection of antigen expression","authors":"Linhua Tian,&nbsp;Aaron R. Nelson,&nbsp;Tyler Lowe,&nbsp;Linda Weaver,&nbsp;Constance Yuan,&nbsp;Hao-Wei Wang,&nbsp;Paul DeRose,&nbsp;Maryalice Stetler-Stevenson,&nbsp;Lili Wang","doi":"10.1002/cyto.b.22155","DOIUrl":"10.1002/cyto.b.22155","url":null,"abstract":"<p>Since response to antigen-based immunotherapy relies upon the level of tumor antigen expression we developed an antigen quantification assay using ABC values. Antigen quantification as a clinical assay requires methods for quality control and for interlaboratory and inter-cytometer platform standardization. A single lot of Cytotrol™ Lyophilized Control Cells (Beckman Coulter) used for all studies. The variability in antigen quantification across 4 different instrument platforms in 2 separate laboratories was evaluated. The effect of the antibody clone utilized, importance of custom 1:1 molar ratio (fluorophore to protein, F/P) verses off-the-shelf antibodies, and QuantiBrite PE calibration verses linearity calibration combined with a single point scale transformation with CD4 as reference were determined. Use of single lot control cells allowed validation of reproducibility between flow cytometer platforms and laboratories and allowed assessment of different antibody lots, cocktail preparation, and different antibody clones. Off the shelf antibody preparations provide reproducible estimates of antigen density, however custom 1:1 unimolar antibody preparations should be utilized for definitive measurement of antigen expression.Geometric Mean fluorescent Intensity (GeoMFI) was not comparable across instruments and inter-laboratory. The use of CD4 as the reference marker can minimize variability in ABC values. Comparable antigen quantification is vital in managing patients receiving antigen-based immunotherapy. If this assay is to be utilized in a clinical setting, quality control methods have to be instituted to assure reproducibility and allow validation across laboratories. We have demonstrated that use of a lyophilized cell control is highly valuable in achieveing these goals.</p>","PeriodicalId":10883,"journal":{"name":"Cytometry Part B: Clinical Cytometry","volume":"106 1","pages":"25-34"},"PeriodicalIF":3.4,"publicationDate":"2024-01-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139461315","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"CD34 and CD117 negative pure erythroid leukemia and phenotypic differences with acute megakaryoblastic leukemia","authors":"Alejandra Altube,&nbsp;Daniela Chelin,&nbsp;Mariela Gomez,&nbsp;Cecilia Malusardi,&nbsp;Dolores Sciaccaluga,&nbsp;Cecilia Cabral,&nbsp;Mariangeles Auat","doi":"10.1002/cyto.b.22160","DOIUrl":"10.1002/cyto.b.22160","url":null,"abstract":"","PeriodicalId":10883,"journal":{"name":"Cytometry Part B: Clinical Cytometry","volume":"106 2","pages":"149-152"},"PeriodicalIF":3.4,"publicationDate":"2024-01-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139377332","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Issue highlights—November 2023","authors":"Professor Alberto Orfao","doi":"10.1002/cyto.b.22154","DOIUrl":"https://doi.org/10.1002/cyto.b.22154","url":null,"abstract":"&lt;p&gt;This new issue of Cytometry B (Clinical Cytometry) consists of five main manuscripts which contain original research in the field of clinical cytometry. A manuscript describing a simple (new) method for preservation of urinary cells for subsequent flow cytometric analyses (Freund et al., &lt;span&gt;2023&lt;/span&gt;) opens this issue of the journal. It is followed by three papers related to the application of flow cytometry in the field of acute leukemias. In the first two manuscripts distinct assays for measurable residual disease (MRD) monitoring in acute myeloblastic leukemia (AML) (Tettero et al., &lt;span&gt;2023&lt;/span&gt;) and B-cell precursor (BCP) acute lymphoblastic leukemia (ALL) (Arunachalam et al., &lt;span&gt;2023&lt;/span&gt;) are (technically and clinically) validated, whereas the third one consists of a comparison and validation of four immunophenotypic scoring systems for the diagnosis of early-T precursor (ETP) acute lymphoblastic leukemia (ALL) (Basavaraju et al., &lt;span&gt;2023&lt;/span&gt;). The fifth article in this issue of Cytometry B revisits the application of flow cytometry for HLA-B27 typing through the comparison of 3 CE-IVD certified methods (Waeckel et al., &lt;span&gt;2023&lt;/span&gt;). Three letters to the editor complete the contents of the November issue of Cytometry B, in which different aspects of three clinically relevant flow cytometric assays for sepsis (Haem-Rahimi et al., &lt;span&gt;2023&lt;/span&gt;), drug-induced hypersensitivity (Ebo et al., &lt;span&gt;2023&lt;/span&gt;) and diagnostic screening of acute leukemias (Axler et al., &lt;span&gt;2023&lt;/span&gt;), are briefly addressed. In this section, I will summarize the contents and highlight the most relevant contributions of the above papers in four separate blocks related to the fields of (i) the flow cytometric analysis of samples with low cell viability, (ii) the flow cytometric diagnosis and monitoring of acute leukemias, (iii) HLA-B27 typing and (iv) the feasibility to measure HLADR expression levels on stabilized blood monocytes and blood circulating drug-specific T cells in the diagnostic work-up of sepsis and drug-hypersensitivity, respectively.&lt;/p&gt;&lt;p&gt;Flow cytometry assays used in diagnostic laboratories have mostly focused on blood samples and to a less extent also, in bone marrow and other tissue and body fluid specimens. Despite the high frequency of kidney and urinary tract diseases in the general population, and the frequent need for invasive diagnostic procedures (e.g., kidney biopsy), urine has been one of the less explored and used specimens among the distinct types of body fluids evaluated in (clinical) flow cytometry. Of note, urine samples are frequently obtained for conventional biochemistry assays, including analysis of proteinuria, and for the evaluation of the urine sediment by conventional, for example, cytomorphology. In contrast, flow cytometric analysis of urinary cells (e.g., immune cells, podocytes or epithelial cells) is rarely used in routine diagnostics, despite it has proven to provide valuable infor","PeriodicalId":10883,"journal":{"name":"Cytometry Part B: Clinical Cytometry","volume":"104 6","pages":"413-416"},"PeriodicalIF":3.4,"publicationDate":"2023-12-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/cyto.b.22154","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138739912","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Editorial on IVD cellular assay validation","authors":"Bruce H. Davis","doi":"10.1002/cyto.b.22156","DOIUrl":"10.1002/cyto.b.22156","url":null,"abstract":"&lt;p&gt;The article in this issue of Cytometry B on “Standardization of Flow Cytometric Detection of Antigen Expression” by the NCI clinical cytometry group formerly headed by Maryalice Stetler-Stevenson and the NIST group headed by Lili Wang is deserving of not only an accompanying editorial, but special attention by all those readers intending to work in clinical cytometry for the coming decade, as it describes an important future component of diagnostic cellular analysis (Tain et al., &lt;span&gt;2023&lt;/span&gt;). Specifically, the ability to measure the antigen (or probe target) expression on well-characterized cell populations will be a vital component of not only monitoring of patients with malignancy, as discussed in the article by Tian et al. herein (Tain et al., &lt;span&gt;2023&lt;/span&gt;) but also monitoring of a variety of immune responses or therapeutically altered defined cell populations. A few predictions as to what true antigen quantitation will provide: (1) treatment of sickle cell disease will be adjusted through a standardized measurement of the level of hemoglobin F in F cells (Hgb F containing RBCs) in order to retard the sickling process (De Souza et al., &lt;span&gt;2023&lt;/span&gt;); (2) patients with severe infection, cytokine storms and certainly sepsis will be monitored for a combination of activation markers (CD64, CD169, HLA-Dr and others) on neutrophils, monocytes and other cell types for informative and actionable changes regarding the immune status (Bourgoin et al., &lt;span&gt;2020&lt;/span&gt;; Davis et al., &lt;span&gt;2006&lt;/span&gt;; Davis &amp; Bigelow, &lt;span&gt;2005&lt;/span&gt;; Ortillon et al., &lt;span&gt;2021&lt;/span&gt;; Schiff et al., &lt;span&gt;1997&lt;/span&gt;); (3) Rapid assays for the genetic expression of newly induced targets (CAR-T cells, adenovirus insertion of other targeting receptors, etc.).&lt;/p&gt;&lt;p&gt;The paper also compares two commonly advocated quantitation methods, PE labeled beads to derive average or median antibody binding capacity (ABC) per cell (Davis et al., &lt;span&gt;1998&lt;/span&gt;) vs. single point transformation or ratiometric comparison of the targeted cell population to the CD4 expression on helper T cells using an assumed 40,000 CD4 mAb binding sites per cell (Degheidy et al., &lt;span&gt;2016&lt;/span&gt;; Wang et al., &lt;span&gt;2016&lt;/span&gt;). Other technical variables the paper convincingly observed is that purified 1:1 PE:antibody preparations give better precision than regular off-the-shelf PE-labeled antibody lots, even if the measured F/P ratio of the off the shelf preparation is close to 1.00. Not surprisingly the study provides quantitative evidence that clone selection does matter and different clones with the same reported target antigen specificity can give variable results, up to nearly a two-fold difference in ABC units and this difference was in no way correctable using the reported F/P ratio of the antibody lot. While the use of 1:1 PE:antibody preparation along with spectrally matched beads for ABC quantitation gave acceptable imprecision with a CV between four instruments","PeriodicalId":10883,"journal":{"name":"Cytometry Part B: Clinical Cytometry","volume":"106 1","pages":"9-10"},"PeriodicalIF":3.4,"publicationDate":"2023-12-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/cyto.b.22156","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138686029","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}