Diagnostic Value of Flow Cytometry Standardized Using the European LeukemiaNet for Myelodysplastic Syndrome

2019 ◽  
Vol 143 (2) ◽  
pp. 140-145 ◽  
Author(s):  
Akihiro Takeuchi ◽  
Osamu Imataki ◽  
Hiroyuki Kubo ◽  
Akihiro Kondo ◽  
Kayoko Seo ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Myeloproliferative Neoplasms ◽  
Diagnostic Value ◽  
Morphological Examination ◽  
Valuable Alternative ◽  
Ringed Sideroblasts ◽  
Hematological Disorders ◽  
Low Incidence ◽  
Low Sensitivity ◽  
Undetermined Significance

Background: Myelodysplastic syndromes (MDS) and idiopathic cytopenia of undetermined significance (ICUS) are heterogeneous hematological disorders characterized by hematopoietic dysplasia and/or chromosomal aberrancy. Objectives: This study aimed to evaluate the diagnostic value of flow cytometry standardized using the European LeukemiaNet (ELN) for MDS and ICUS by analyzing samples obtained from patients with cytopenia based on morphological examination, cytogenetic analysis, and flow cytometry. Methods: We retrospectively analyzed bone marrow samples aspirated from 253 consecutive patients (median age: 66 years [range: 1–92]) to identify the cause of cytopenia. Results: Sixty patients presented with MDS, and 16 with ICUS. MDS subtypes were distributed as follows: MDS with single-lineage dysplasia (n = 10); MDS with multi-lineage dysplasia (n = 10); MDS with ringed sideroblasts (n = 4); MDS with excess blasts-1 (n = 9); MDS with excess blasts-2 (n = 13), MDS unclassified (n = 5); 5q-syndrome (n = 6); and MDS/myeloproliferative neoplasms (n = 3). Four representative ELN indexes were used. Two or more ELN MDS indexes were in the abnormal range in 35 MDS cases (58.3%) and 4 ICUS cases (25.0%). Conclusions: Morphological examination remains the standard for MDS diagnosis. Considering the low incidence of genetically proven ICUS (20.2–27.5%), the low sensitivity of ELN MDS indexes for ICUS is considered a valuable alternative.

scholarly journals Comparing the Diagnostic Value of Serum D-Dimer to CRP and IL-6 in the Diagnosis of Chronic Prosthetic Joint Infection

2020 ◽  
Vol 9 (9) ◽  
pp. 2917
Author(s):  
Thomas Ackmann ◽  
Burkhard Möllenbeck ◽  
Georg Gosheger ◽  
Jan Schwarze ◽  
Tom Schmidt-Braekling ◽  
...  
Keyword(s):  
Sensitivity And Specificity ◽  
Joint Infection ◽  
Revision Arthroplasty ◽  
Preoperative Assessment ◽  
Diagnostic Value ◽  
Musculoskeletal Infection ◽  
Prosthetic Joint ◽  
The Individual ◽  
Low Sensitivity ◽  
D Dimer

Introduction: D-dimer is a diagnostic criterion for periprosthetic joint infection (PJI) of the Musculoskeletal Infection Society (MSIS) in 2018. The aim of this study was to evaluate the serum D-dimer values in comparison to C-reactive protein (CRP) and interleukin-6 (IL-6) for the diagnosis of PJI. Materials and Methods: We included 119 patients (50 women, 69 men; 71 knees, 48 hips) undergoing revision arthroplasty with preoperative assessment of CRP, IL-6, and serum D-dimer. Cases were classified as infected or aseptic based on the MSIS criteria of 2018. Receiver operating curves and Youden’s index were used to define an ideal cut-off value and sensitivity and specificity for the individual parameters, and respective combinations were calculated using cross-tables. Results: The median D-dimer level (2320 vs. 1105 ng/mL; p < 0.001), the median CRP level (4.0 vs. 0.5 mg/dL; p < 0.001), and the median IL-6 level (21.0 vs. 5.0 pg/mL; p < 0.001) were significantly higher in the group of PJI compared to the group with aseptic failure. The calculated optimal cut-off values were 2750 ng/mL (AUC 0.767) for D-dimer, 1.2 mg/dL (AUC 0.914) for CRP, and 10.0 pg/mL (AUC 0.849) for IL-6. D-dimer showed a sensitivity of 38% and specificity of 94%, whereas the CRP and IL-6 had sensitivities of 88% and 76%, and specificities of 87% and 92%, respectively. Conclusion: In comparison with CRP and IL-6, serum D-dimer showed low sensitivity and specificity in our cohort. While CRP and IL-6 combination had the highest sensitivity, a combination of Il-6 and D-dimer or CRP and IL-6 had the highest specificity.

scholarly journals Comparison of a Classical Phagocytosis Assay and a Flow Cytometry Assay for Assessment of the Phagocytic Capacity of Sera from Adults Vaccinated with a Pneumococcal Conjugate Vaccine

2001 ◽  
Vol 8 (2) ◽  
pp. 245-250 ◽  
Author(s):  
Wouter T. M. Jansen ◽  
Merja Väkeväinen-Anttila ◽  
Helena Käyhty ◽  
Moon Nahm ◽  
N. Bakker ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Defense Mechanism ◽  
Worker Safety ◽  
Enzyme Linked Immunosorbent Assay ◽  
Phagocytosis Assay ◽  
Phagocytic Capacity ◽  
Killing Assay ◽  
Potential Efficacy ◽  
Antibody Titers ◽  
Low Sensitivity

ABSTRACT Antibody- and complement-mediated phagocytosis is the main defense mechanism against Streptococcus pneumoniae. A standardized, easy to perform phagocytosis assay for pneumococci would be a great asset for the evaluation of the potential efficacy of (experimental) pneumococcal vaccines. Such an assay could replace the laborious phagocytosis assay of viable pneumococci (classical killing assay). Therefore, a newly developed phagocytosis assay based on flow cytometry (flow assay) was compared with the conventional killing assay and enzyme-linked immunosorbent assay (ELISA), using sera obtained from adults pre- and postvaccination with either a bivalent conjugate, a tetravalent conjugate, or the 23-valent polysaccharide vaccine. Highly significant correlations were observed between flow assay phagocytosis titers, killing assay phagocytosis titers, and ELISA antibody titers for serotype 6B and 23F as well. For serotype 19F, strong correlations were only observed between killing assay and ELISA titers. A potential drawback of the flow assay might be the low sensitivity compared with that of the killing assay. The choice of what assay to use, however, will depend on the objectives of the assay. When speed, easy performance, sample throughput, improved worker safety, absence of influence of antibiotics, and absence of false positives are the major criteria, the flow assay is the method of choice. When higher sensitivity is the major requirement, the classical killing assay should be used.

scholarly journals Endoscopic Biopsy in Differential Diagnosis of Colorectal Serrated Lesions

2020 ◽  
Vol 30 (3) ◽  
pp. 42-48
Author(s):  
K. D. Khalin ◽  
M. Yu. Agapov ◽  
L. V. Zvereva ◽  
K. V. Stegniy
Keyword(s):  
Differential Diagnosis ◽  
Endoscopic Resection ◽  
Tumour Size ◽  
Endoscopic Biopsy ◽  
Morphological Examination ◽  
Serrated Lesions ◽  
Low Sensitivity ◽  
Average Size ◽  
Sensitivity Specificity ◽  
Serrated Adenomas

Background. Preoperative biopsy is recommended for morphological verification of colorectal epithelial neoplasms prior to their endoscopic resection. However, histological reports for endoscopic biopsy and resected lesions are not reliably consistent.Aim. Assessment of sensitivity, specificity and accuracy of endoscopic biopsy in differential diagnosis of colorectal serrated adenomas and risk factors for variance between biopsy results and morphological examination of completely resected lesions.Materials and methods. The assay used data on 56 morphologically verified serrated adenomas diagnosed and resected in 50 patients (14 men, 36 women; average age 66.9 ± 10.5 years). Biopsy was taken from all tumours before endoscopic resection. Results of morphological examination of biopsy samples and resected tumours were analysed and compared. Sensitivity, specificity and accuracy of biopsy was assessed, with the tumour size and type and biopsy forceps system taken as criteria.Results. The identified cases included 22 (39.3%) right-colon, 21 (37.5%) left-colon and 13 (23.2%) rectal lesions of 28.5 ± 2.6 mm average size. Polypoid were 17 (30.3%), non-polypoid — 6 (10.7%) and spreading — 33 (59%) of the tumours. Full consistency of morphological examination was observed for 12 cases (21.4%). In 9 cases (16%), dysplasia was established as mild-graded with biopsy, whilst the eradicated tumours contained severe dysplastic foci. Foci of adenocarcinoma were detected in 10 tumours (including 2 with submucosal invasion), but only 2 cases were correctly diagnosed for malignant adenoma with biopsy. Tumour morphology was misidentified in 32 cases (57.1%).Conclusions. Preoperative forceps biopsy is shown to possess low sensitivity in differential diagnosis of serrated colorectal lesions and very low sensitivity to predict malignant serrated adenomas.

scholarly journals The Rule of Histology in the Diagnosis of Periprosthetic Infection: Specific Granulocyte Counting Methods and New Immunohistologic Staining Techniques may Increase the Diagnostic Value

2016 ◽  
Vol 10 (1) ◽  
pp. 457-465 ◽  
Author(s):  
Friedrich Boettner ◽  
Gabriele Koehler ◽  
Alexander Wegner ◽  
Tom Schmidt-Braekling ◽  
Georg Gosheger ◽  
...  
Keyword(s):  
Periprosthetic Infection ◽  
High Specificity ◽  
Diagnostic Value ◽  
Lower Sensitivity ◽  
Specific Staining ◽  
Tissue Samples ◽  
Hip And Knee Arthroplasty ◽  
Staining Techniques ◽  
Low Sensitivity ◽  
Intraoperative Test

Objective: The current study investigates the diagnostic accuracy of the criteria described for frozen sections and whether modern leukocyte specific staining techniques including leukocyte peroxidase and Naphtol-AS-D-chloroacetate-esterase will improve the accuracy of the intra-operative histology. Method: 77 patients undergoing revision total hip and knee arthroplasty were included in this retrospective study. Patients were grouped into septic and aseptic based on intraoperative cultures. Tissue samples were analyzed utilizing the Mirra, Feldman, Lonner, Banit and Athanasou criteria. Results: An experienced pathologist had a high specificity (96%), but rather low sensitivity (57%) diagnosing infection. By using the Banit-, Mirra-, or Athanasou-criteria the sensitivity is increased to 0.90. The Feldman- and Lonner-criteria have a lower sensitivity (0.48 and 0.38), however, an increased specificity of 0.96 and 0.98, respectively. The Banit cut off has the highest accuracy (86%). MPOX and NACE staining increased the sensitivity and accuracy up to 100% and 92% respectively. Conclusion: Banit’s cut off is the most accurate histologic criteria to diagnose infection. Modern leukocyte specific staining techniques slightly improve the accuracy. The synovial fluid white blood cell count appears to be the most accurate intraoperative test.

Mutant CXCR4 Identified in Neutropenic Patients with Myelokathexis Impairs Survival of Human Myeloid Cells.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3071-3071
Author(s):  
Vahagn Makaryan ◽  
David C. Dale ◽  
Andrew A. Aprikyan
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Cell Survival ◽  
Mutational Analysis ◽  
Bone Marrow Cells ◽  
Myeloid Cells ◽  
Receptor Internalization ◽  
Morphological Examination ◽  
Exon 2 ◽  
Whim Syndrome

Abstract Myelokathexis (WHIM syndrome) is a very rare hematopoietic congenital disorder that is characterized by extremely low level of circulating neutrophils in peripheral blood. It is inherited as an autosomal dominant disease and is diagnosed in early childhood. These patients may have hypogammaglobulinemia and suffer from recurrent infections associated with warts. The hallmark of myelokathexis is a hyperplastic bone marrow and hypersegmented neutrophils with nuclear lobs connected with thin filaments. Myelokathexis is due to a characteristic retention of mature neutrophils in bone marrow, which are not being released to peripheral circulation. We and others reported abnormal cell survival characteristics and impaired bcl-x expression in bone marrow myeloid cells of myelokathexis patients that was partially restored by G-CSF treatment. Recently, it has also been reported that heterozygous truncation mutations in the carboxyterminal domain of the CXCR4 gene, a sole receptor for SDF-1 chemokine, were observed in most, but not all of the families with WHIM syndrome. Subsequently, an impaired receptor internalization and increased chemotaxis towards SDF-1 have been observed in cells expressing truncated CXCR4. Nevertheless, the mechanism of mutant CXCR4 induced myelokathexis remains largely unknown. We performed mutational analysis of the CXCR4 gene in 3 unrelated families with myelokathexis and identified a previously reported R334ter truncation mutation in exon 2 in two of the families. In addition, two silent polymorphisms have been identified in exon 2 of the CXCR4 gene in one of these patients. The third family with afflicted mother and son had a new mutation in the CXCR4 carboxyterminal domain, which resulted in deletion of the last 16 amino acids and subsequent frame shift. None of these mutations were observed in healthy volunteers examined. Since the morphological examination by electron microscopy and flow cytometry analysis of bone marrow cells from some of these patients revealed characteristic apoptotic features, we examined the effect of mutant CXCR4 gene expression on survival of human promyelocytic HL-60 cells. Preliminary data demonstrated that human promyelocytic cells transfected with truncated CXCR4 exhibited impaired cell survival characteristics compared with control HL-60 cells transfected with intact CXCR4. The truncated, but not wild type CXCR4 also increased apoptosis in HL-60 cells induced to differentiate along the granulocytic pathway as determined by flow cytometry of annexin V labeled cells. Thus, these data link together the abnormal survival of proliferating and differentiating myeloid cells in WHIM syndrome with mutant CXCR4 expression. Current studies are focused on elucidation of specific signaling pathways mediating mutant CXCR4-triggered myelokathexis.

Detection of Circulating Antigen-Specific CD8+ Lymphocytes against Leukemia Associated Antigens WT1, PR1 and Survivin Using HLA 0201 Pentamers After Allogeneic Stem Cell Transplantation.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 5115-5115
Author(s):  
Juana Serrano-Lopez ◽  
Joaquin Sanchez Garcia ◽  
Josefina Serrano ◽  
Carmen Martin ◽  
Rafael Rojas ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Stem Cell ◽  
Stem Cell Transplantation ◽  
Peripheral Blood ◽  
Median Percentage ◽  
Blood Samples ◽  
Hematological Disorders ◽  
Multiparametric Flow Cytometry ◽  
Leukemia Antigen

Abstract Abstract 5115 INTRODUCTION Allogeneic stem cell transplantation (allo-SCT) is a potentially curative treatment option for patients with hematological disorders. Alloreactive donor-derived T lymphocytes exert a beneficial graft-versus-leukemia (GVL) effect through the recognition of leukemia-restricted (or preferentially expressed) antigens as Wilms tumor protein (WT1), survivin (SURV) or proteinase (PR1). Currently research in transplant immunology focuses in enhancing GVL while preventing the deleterious graft-versus-host disease (GVHD) that could be achieved by manipulating donor-derived antigen-specific T-populations. In this study we tested the presence of peripheral blood leukemia-associated antigen-specific CD8+ T-lymphocytes during post allo-SCT follow-up. PATIENTS AND METHODS Forty-three consecutive HLA*0201 patients (homo or heterozygotous) undergoing conventional myeloablative (n=24) or non-myeloablative (n=19) allo-SCT as treatment of hematological disorders were included. Allogeneic donor was an HLA-identical sibling in 26 cases (60.5%) and unrelated in 17 cases (39.5%). Hematopoietic stem cell source included mobilized peripheral blood (n=20), bone marrow (n=18) and umbilical cord blood (n=5). As GVHD prophylaxis regimens Cyclosporine plus Methotrexate (n=20) or Cyclosporine plus Mofetil micofenolate (n=23) were employed. In addition, 22 patients received rabbit antithymocyte globulin at 6-8mg/kg. At last follow-up four patients had relapsed 9-14 months after allo-SCT. We sought for leukemia-antigen specific CD8 lymphocytes in peripheral blood samples drawn within a median of 7 months (range 2-38) when lymphocyte recovery had occurred and complete donor chimerism was achieved. We used four color multiparametric flow cytometry in a FACSCanto II acquiring at least 5 ×105 viable (Propidium Iodide low) lymphoid gated events, stained with MnAbs: CD8-FITC and CD3PE/APC MnAb. To identify leukemia-antigen specific CD8 lymphocytes we used class I HLA pentamers 0201 APC or PE conjugated (Proimmune, London, UK) against the following nonapeptides: Proteinase 1: VLQELNVTV (169-177) WT1: RMFPNAPYL (126-134) and SURV: ELTLGEFLKL (95-104). As positive staining control we used CMV pp65: NLVPMVATV (495-503) and as negative controls we used irrelevant nonapeptide and peripheral blood samples from patients lacking HLA* 0201 genotype. RESULTS Detection of donor-derived CD8+ lymphocytes against CMV pp65 occurred in 61% of recruited patients with a median percentage of 0.1% (range 0.03-13 over CD3+CD8+ events). Likewise, it was possible to detect CD8+ lymphocytes specific for PR1, WT1 and SURV in 65.2%, 47.8% y 39.1% of recruited patients respectively. Median percentage of PR1 and WT1 leukemia-antigen specific lymphocytes was 0.1% (range 0.04-1% over CD3+CD8+ events) and for WT1 0.1% (range: 0.01-0.2%). Detection of leukemia-antigen specific CD8+ lymphocytes was not significantly associated with clinical variables such as conditioning regimen (conventional or non-myeloablative), Disease status at transplant, donor type (sibling or unrelated), ATG use or HLA-disparity degree. The presence of WT1 specific CD8+ lymphocytes was significantly more frequent in patients undergoing allo-SCT for lymphoid hematological malignancy (p=0.04). By contrast, the presence of circulating anti-PR1 specific CD8+ lymphocytes was not more frequently found in patients undergoing allo-SCT for myeloid malignancies. Of note, none of the four patients who eventually relapsed harbored circulating leukemia-specific CD8+ lymphocytes. CONCLUSIONS Multiparametric flow cytometry is a useful tool to detect and quantify rare donor-derived CD8+ lymphocytes specific for leukemia-associated antigens as PR1, WT1 or SURV. The presence of these populations in peripheral blood is not associated to conventional clinical variables and in our series anti-WT1 CD8+ lymphocytes were more frequently detected in patients receiving allo-SCT for lymphoid malingnacies. By contrast, larger series are needed to assess if the lack of these leukemia-associated antigen-specific CD8 lymphocytes in peripheral blood could identify patients in a higher risk of relapse. Financial support This study was supported by a grant of Conserjeria de Salud, Junta de Andalucia 2006/0355. J. Serrano López is a post-doc fellow from Fundación Española de Hematología y Hemoterapia Disclosures No relevant conflicts of interest to declare.

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