Publications by authors named "Jolene A Bradford"

Even though anti-PD-1/PD-L1 immune checkpoint inhibitors (ICIs) in non-small cell lung cancer (NSCLC) have improved survival, a high percentage of patients still do not respond to ICIs. Myeloid-derived suppressor cells (MDSCs) are circulating cells that express PD-L1 and can infiltrate and proliferate in the tumor microenvironment, inducing immunosuppression. By evaluating changes in PD-L1 expression of live peripheral blood MDSCs, we are able to define a new PD-L1 index, useful in predicting ICI escape in NSCLC patients before initiating anti-PD-1/PD-L1 immunotherapy.

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Leukemic stem cells (LSCs) possess similar characteristics to normal hematopoietic stem cells, including self-renewal capacity, quiescence, ability to initiate leukemia, and drug resistance. These cells play a significant role in leukemia relapse, persisting even after apparent remission. LSCs were first described in 1994 by Lapidot et al.

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  • The study evaluates a new volumetric counting method for CD34+ cells in blood samples, which aims to improve accuracy over traditional methods that use reference microbeads.
  • The issue with the "vanishing counting bead phenomenon" affects the reliability of results obtained with bead-based systems but has been addressed through the development of a true volumetric counting system.
  • Results showed no significant difference in CD34+ cell counts between volumetric and non-volumetric methods, suggesting that the new volumetric technique could simplify sample preparation and reduce analysis bias.
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The programmed cell death protein 1/programmed cell death protein ligand 1 (PD-1/PD-L1) axis is one of the most widely recognized targets for cancer immunotherapy. Importantly, PD-L1 conformational changes can hinder target binding when living cells are used. Antibody affinity, equilibrium binding, association and dissociation rates, and other affinity-related constants are fundamental to ensure target saturation.

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Plastic pollution is a global problem. Animals and humans can ingest and inhale plastic particles, with uncertain health consequences. Nanoplastics (NPs) are particles ranging from 1 nm to 1000 nm that result from the erosion or breakage of larger plastic debris, and can be highly polydisperse in physical properties and heterogeneous in composition.

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Flow cytometry (FCM) enumeration of peripheral blood dendritic cells (PBDCs) is a minimally invasive procedure extremely useful for immunological studies. Numbers of PBDCs vary depending on age, lifestyle, or in pathologies like cancer, leukemia or immunodeficiencies. Conventional methods for PBDC identification by FCM involve red blood cell lysis using either formaldehyde or ammonium chloride-based solutions.

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Doublet discrimination is usually based on pulse analysis of light scatter parameters. A combination of two pulse parameters (Area, A; Height, H; or Width, W) can be used to discriminate a pulse originated in a single cell from a pulse originated from cells stuck together. Fluorescence signals can be also used to discriminate aggregates, being essential to identify cells in the G2/M phase from doublets in the G0/G1 phase in cell cycle/DNA applications.

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The challenges associated with analyzing rare cells are dependent on a series of factors, which usually require large numbers of cells per sample for successful resolution. Among these is determining the minimum number of total events needed to be acquired as defined by the expected frequency of the target cell population. The choice of markers that identify the target population, as well as the event rate and the number of aborted events/second, will also determine the statistically significant detection of rare cell events.

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This protocol provides instructions to improve flow cytometry analysis of marrow/peripheral blood cells by avoiding erythrolytic solutions, density gradients, and washing steps. We describe two basic approaches for identifying cell surface antigens with minimal sample perturbation, which have been successfully used to identify healthy and pathologically rare cells. The greatest advantage of these approaches is that they minimize the unwanted effect caused by sample preparation, allowing for improved study of live cells at the point of analysis.

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Current methods for the determination of cell-mediated cytotoxic activity in blood samples usually isolate peripheral blood mononuclear cells by density gradient centrifugation or alternatively use erythrocyte lysis. Both centrifugation and red cell lysis can cause cellular depletion and cell dysfunction, resulting in erroneous measurements. To address limitations of current assays, we developed an improved strategy to determine cellular cytotoxicity using flow cytometry.

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• PD-L1 displays variation of spatial conformation in response to phorbol ester stimulation, which may confer a critical enhancement in binding affinity. • PD-L1 conformational change may be associated with an immunoregulatory mechanism that affects therapies targeting the PD-1/PD-L1 checkpoint. • Eliminating MDSCs by promoting PD-L1 stabilized unfolded states on both PMN- and M-MDSCs could improve immunotherapy efficacy.

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For several decades, cell-mediated cytotoxicity has been measured using the Cr release assay. This assay, however, has several drawbacks and flow cytometry has been used as an alternative to measure cytotoxic activity. Here, we present a quantitative method for cell-mediated cytotoxicity studies, preserving cellular function with minimal sample manipulation.

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Translational research has improved the diagnosis and follow-up of hematological diseases and malignancies. However, some classical diagnostics used for research and clinical practice that have remain practically unchanged for decades may be better addressed through advances in flow cytometry technology, whereby more precise measurements may be implemented in a straightforward manner. The current method for semiquantitative analysis of granulocytic alkaline phosphatase (GAP) activity is still based on observer-dependent color-intensity classification.

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In this prospective hospital-based cohort study that included 43 newly diagnosed patients with acute myeloid leukemia, flow cytometric cellular alkaline phosphatase (ALP) activity within primitive leukemic cells allowed us to identify two groups of patients at diagnosis according to the numbers of leukemic blasts expressing ≥ 12% of ALP+ cells (27 patients, Group A) and less than 12% of ALP+ cells (16 patients, Group B). Differences in outcome for complete response, relapse or treatment resistance, and exitus were statistically analyzed and were significant, when comparing the two groups. The overall survival (OS) and event-free survival (EFS) differences between Group A and B were statistically significant.

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These guidelines are a consensus work of a considerable number of members of the immunology and flow cytometry community. They provide the theory and key practical aspects of flow cytometry enabling immunologists to avoid the common errors that often undermine immunological data. Notably, there are comprehensive sections of all major immune cell types with helpful Tables detailing phenotypes in murine and human cells.

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Distortions of the normal bi-concave disc shape for red blood cells (RBCs) appear in a number of pathologies resulting from defects in cell membrane skeletal architecture, erythrocyte ageing, and mechanical damage. We present here the potential of acoustic cytometry for developing new approaches to light-scattering based evaluation of red blood cell disorders and of the effects of storage and ageing on changes or damage to RBCs membranes. These approaches could be used to immediately evaluate the quality of erythrocytes prior to blood donation and following transfusion.

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Red blood cell lysis is an integral part of many flow cytometry protocols. It's potential to cause artifacts has been known for decades, but lysis free sample preparation has failed to replace lysis in most applications. Studies of various lysing protocols on cell losses and effects on phenotypic markers and cell function began early in the history of immunophenotyping and continue to this day.

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The measurement of cellular proliferation is fundamental to the assessment of cellular health, genotoxicity, and the evaluation of drug efficacy. Labeling, detection, and quantification of cells in the synthesis phase of cell cycle progression are not only important for characterizing basic biology, but also in defining cellular responses to drug treatments. Changes in DNA replication during S-phase can provide valuable insights into mechanisms of cell growth, cell cycle kinetics, and cytotoxicity.

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Article Synopsis
  • - Adult human mesenchymal stem cells (hMSC) are unique cells found mainly in bone marrow and fat that can turn into various tissue types like bone and cartilage.
  • - The International Society for Cellular Therapy (ISCT) has set criteria to define hMSC, including their ability to adhere to surfaces and differentiate into specific cell types.
  • - Flow cytometry is a key technique used for analyzing hMSC by measuring their immunophenotypic profile, growth potential, and DNA content, allowing researchers to make better decisions regarding cell expansion and harvesting.
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  • Changes in DNA replication during S-phase provide key insights into cell growth, cycle timing, and how cells react to toxins.
  • A popular method to track cell proliferation involves using thymidine analogs, where multiple analogs at different times help map cell cycle kinetics.
  • This text introduces a dual-pulse method combining the thymidine analog EdU with BrdU, allowing precise detection of cell division without cross-reactivity between the two methods.
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  • The study explores the use of EdU (5-ethynyl-2'-deoxyuridine) as an alternative to BrdU (5-bromo-2'-deoxyuridine) for assessing cell proliferation in breast cancer cells.
  • The researchers found that EdU allows for accurate continuous cell labeling and comparable results to BrdU, but long-term exposure to EdU can negatively impact cell viability, particularly in SK-BR-3 cells.
  • While both SK-BR-3 and BT474 cells showed cell cycle arrests with EdU treatment, only SK-BR-3 experienced necrotic cell death, making it important to evaluate individual cell response to EdU when testing cell proliferation.
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Background: Conventional immuno-based multiparameter flow cytometric analysis has been limited by the requirement of a dedicated detection channel for each antibody-fluorophore set. To address the need to resolve multiple biological targets simultaneously, flow cytometers with as many as 10-15 detection channels have been developed. In this study, a new Zenon immunolabeling technology is developed that allows for multiple antigen detection per detection channel using a single fluorophore, through a unique method of fluorescence-intensity multiplexing.

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