Neutrophils are the most abundant peripheral immune cells and thus, are continually replenished by bone marrow-derived progenitors. Still, how newly identified neutrophil subsets fit into the bone marrow neutrophil lineage remains unclear. Here, we use mass cytometry to show that two recently defined human neutrophil progenitor populations contain a homogeneous progenitor subset we term "early neutrophil progenitors" (eNePs) (Lin-CD66b+CD117+CD71+). Surface marker- and RNA-expression analyses, together with in vitro colony formation and in vivo adoptive humanized mouse transfers, indicate that eNePs are the earliest human neutrophil progenitors. Furthermore, we identified CD71 as a marker associated with the earliest neutrophil developmental stages. Expression of CD71 marks proliferating neutrophils, which were expanded in the blood of melanoma patients and detectable in blood and tumors from lung cancer patients. In summary, we establish CD117+CD71+ eNeP as the inceptive human neutrophil progenitor and propose a refined model of the neutrophil developmental lineage in bone marrow.
Copyright © 2020 Elsevier Inc. All rights reserved.

DotScan antibody microarrays were initially developed for the extensive surface profiling of live leukemia and lymphoma cells. DotScan's diagnostic capability was validated with an extensive clinical trial using mononuclear cells from the blood or bone marrow of leukemia or lymphoma patients. DotScan has also been used for the profiling of surface proteins on peripheral blood mononuclear cells (PBMC) from patients with HIV, liver disease, and stable and progressive B-cell chronic lymphocytic leukemia (CLL). Fluorescence multiplexing allowed the simultaneous profiling of cancer cells and leukocytes from disaggregated colorectal and melanoma tumor biopsies after capture on DotScan. In this chapter, we have used DotScan for the surface profiling of extracellular vesicles (EV) recovered from conditioned growth medium of cancer cell lines and the blood of patients with CLL. The detection of captured EV was performed by enhanced chemiluminescence (ECL) using biotinylated antibodies that recognized antigens expressed on the surface of the EV subset of interest. DotScan was also used to profile EV from the blood of healthy individuals and the ascites fluid of ovarian cancer patients. DotScan binding patterns of EV from human plasma and other body fluids may yield diagnostic or prognostic signatures for monitoring the incidence, treatment, and progression of cancers.

The B cell antigen receptor (BCR) plays an essential role in all phases of B cell development. Here we show that the extracellular domains of murine and human Igbeta form an I-set immunoglobulin-like structure with an interchain disulfide between cysteines on their G strands. Structural and sequence analysis suggests that Igalpha displays a similar fold as Igbeta. An Igalphabeta heterodimer model was generated based on the unique disulfide-bonded Igbeta dimer. Solution binding studies showed that the extracellular domains of Igalphabeta preferentially recognize the constant region of BCR with mu chain specificity, suggesting a role for Igalphabeta to enhance BCRmu chain signaling. Cluster mutations on Igalpha, Igbeta, and a membrane-bound form of immunoglobulin (mIgM) based on the structural model identified distinct areas of potential contacts involving charged residues on both subunits of the coreceptor and the Cmu4 domain of mIgM. These studies provide the first structural model for understanding BCR function.
Copyright 2010 Elsevier Ltd. All rights reserved.

The B-cell receptor (BCR) for antigen is composed of surface immunoglobulin (sIg), which provides antigen specificity, and a noncovalently associated signaling unit, the CD79a/b heterodimer. Defects in CD79 can influence both BCR expression and signaling and may explain why cells from certain malignancies, such as B-chronic lymphocytic leukemia (B-CLL), often express diminished and inactive BCR. Recently, an alternative transcript of CD79b (DeltaCD79b) has been reported that is up-regulated in B-CLL and may explain this diminished BCR expression. Here we assess the expression of DeltaCD79b in B-CLL and other lymphoid malignancies and investigate its function. High relative expression of DeltaCD79b was confirmed in most cases of B-CLL and found in 6 of 6 cases of splenic lymphomas with villous lymphocytes (SLVLs) and hairy cell leukemia. In a range of Burkitt lymphoma cell lines, expression of DeltaCD79b was relatively low but correlated inversely with the ability of the BCR to signal apoptosis when cross-linked by antibody (Ab). Interestingly, when Ramos-EHRB cells, which express low DeltaCD79b, were transfected with this transcript, they were transformed from being sensitive to anti-Fcmu-induced apoptosis to being highly resistant. Although DeltaCD79b was expressed as protein, its overexpression did not reduce the level of cell surface BCR. Finally, we showed that the inhibitory activity of DeltaCD79b depended on an intact leader sequence to ensure endoplasmic reticulum (ER) trafficking and a functional signaling immunoreceptor tyrosine-based activation motif (ITAM) in its cytoplasmic tail. These results point to DeltaCD79b being a powerful modulator of BCR signaling that may play an important role in normal and malignant B cells.