The CD33xCD123xCD70 Multispecific CD3-Engaging DARPin MP0533 Induces Selective T Cell-Mediated Killing of AML Leukemic Stem Cells.

The prognosis of patients with acute myeloid leukemia (AML) is limited, especially for elderly or unfit patients not eligible for hematopoietic stem cell (HSC) transplantation. The disease is driven by leukemic stem cells (LSCs), which are characterized by clonal heterogeneity and resistance to conventional therapy. These cells are therefore believed to be a major cause of progression and relapse.

A Multispecific Anti-CD40 DARPin Construct Induces Tumor-Selective CD40 Activation and Tumor Regression.

The CD40 receptor is an attractive target for cancer immunotherapy. Although a modest pharmacodynamic effect is seen in patients following administration of CD40-targeting monoclonal antibodies (mAb), the doses that could be safely administered do not result in a meaningful clinical response, most likely due to the limited therapeutic window associated with systemic CD40 activation. To overcome this issue, we developed a multispecific DARPin construct, α-FAPxCD40, which has conditional activity at the site of disease.

Comparison of Volumetric and Bead-Based Counting of CD34 Cells by Single-Platform Flow Cytometry.

Background: Over 2,000 people a year in the United Kingdom need a bone marrow or blood stem cell transplant. It is important to accurately quantify the hematopoietic stem cells to predict whether the transplant will be successful in replenishing the immune system. However, they are present at low frequency, which complicates accurate quantification.

Rapid comparative immunophenotyping of human mesenchymal stromal cells by a modified fluorescent cell barcoding flow cytometric assay.

Flow cytometry immunophenotyping is a sensitive technique allowing rapid characterization of single cells within heterogeneous populations, but it includes several subjective steps during sample analysis that impact the development of standardized methodology. Proposed strategies to overcome these limitations include fluorescent cell barcoding (FCB), which facilitates multiplexed sample evaluation with increased data reproducibility whilst reducing labeling variation, materials, and time. To date, the FCB assay has found utility for analyzing the phosphorylation status of intracellular targets but has not been intensively employed for cellular immunophenotypic analyses using cell surface markers.

Rapid phenotypic fingerprinting of cell products by robust measurement of ubiquitous surface markers.

Flow cytometry is one of the most versatile and powerful approach for the quantitative analysis of cell suspensions. With widespread applications in basic and clinical research, its commonest use is in the detection of cell populations labelled against markers specific for a particular phenotype. In this study, we aimed to expand the potential of flow cytometry by describing a method based on robust automated quantification of ubiquitous cell surface markers.

Quantification of cells with specific phenotypes I: determination of CD4+ cell count per microliter in reconstituted lyophilized human PBMC prelabeled with anti-CD4 FITC antibody.

A surface-labeled lyophilized lymphocyte (sLL) preparation has been developed using human peripheral blood mononuclear cells prelabeled with a fluorescein isothiocyanate conjugated anti-CD4 monoclonal antibody. The sLL preparation is intended to be used as a reference material for CD4+ cell counting including the development of higher order reference measurement procedures and has been evaluated in the pilot study CCQM-P102. This study was conducted across 16 laboratories from eight countries to assess the ability of participants to quantify the CD4+ cell count of this reference material and to document cross-laboratory variability plus associated measurement uncertainties.

Dynamic monitoring of metal oxide nanoparticle toxicity by label free impedance sensing.

The increased use of nanoparticles in industrial and medical products is driving the need for accurate, high throughput in vitro testing procedures to screen new particles for potential toxicity. While approaches using standard viability assays have been widely used, there have been increased reports of the interactions of nanoparticles with their soluble labels or optical readouts which raise concerns over the potential generation of false positive results. Here, we describe the use of an impedance spectroscopy approach to provide real-time reagent free detection of toxicity for a panel of metal oxide nanoparticles (ZnO, CuO, and TiO(2)).

The tumour-associated antigen L6 (L6-Ag) is recruited to the tetraspanin-enriched microdomains: implication for tumour cell motility.

Tumour-associated antigen L6 (L6-Ag, also known as TM4SF1) regulates tumour cell motility and invasiveness. We found that L6-Ag is abundant on the plasma membrane and on intracellular vesicles, on which it is co-localised with the markers for late endosomal/lysosomal compartments, including Lamp1/Lamp2 proteins and LBPA. Antibody internalisation and live-imaging experiments suggested that L6-Ag is targeted to late endocytic organelles (LEO) predominantly via a biosynthetic pathway.