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Influenza D virus (IDV) was first identified from swine exhibiting influenza-like symptoms in 2011 in the United States (1). IDV RNA or antibodies have been detected worldwide in several animal [...]

Some Plasmodium falciparum repetitive interspersed families of polypeptides (RIFINs)--variant surface antigens that are expressed on infected erythrocytes.sup.1--bind to the inhibitory receptor LAIR1, and insertion of DNA that encodes LAIR1 into immunoglobulin genes generates RIFIN-specific antibodies.sup.2,3. Here we address the general relevance of this finding by searching for antibodies that incorporate LILRB1, another inhibitory receptor that binds to [beta]2 microglobulin and RIFINs through their apical domains.sup.4,5. By screening plasma from a cohort of donors from Mali, we identified individuals with LILRB1-containing antibodies. B cell clones isolated from three donors showed large DNA insertions in the switch region that encodes non-apical LILRB1 extracellular domain 3 and 4 (D3D4) or D3 alone in the variable-constant (VH-CH1) elbow. Through mass spectrometry and binding assays, we identified a large set of RIFINs that bind to LILRB1 D3. Crystal and cryo-electron microscopy structures of a RIFIN in complex with either LILRB1 D3D4 or a D3D4-containing antibody Fab revealed a mode of RIFIN-LILRB1 D3 interaction that is similar to that of RIFIN-LAIR1. The Fab showed an unconventional triangular architecture with the inserted LILRB1 domains opening up the VH-CH1 elbow without affecting VH-VL or CH1-CL pairing. Collectively, these findings show that RIFINs bind to LILRB1 through D3 and illustrate, with a naturally selected example, the general principle of creating novel antibodies by inserting receptor domains into the VH-CH1 elbow. Plasmodium antigens called RIFINs bind to specific antibodies that incorporate the inhibitory receptor LILRB1 through its D3 domain, illustrating the principle of receptor-containing antibodies., Author(s): Yiwei Chen [sup.1] [sup.2] , Kai Xu [sup.3] , Luca Piccoli [sup.1] , Mathilde Foglierini [sup.1] [sup.4] , Joshua Tan [sup.1] , Wenjie Jin [sup.1] [sup.2] , Jason Gorman [...]

Neutralizing antibodies develop in asymptomatic persons with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection; however, the initial immune response is not as strong as in patients with more severe [...]

A considerable fraction of B cells recognize severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) with germline-encoded elements of their B cell receptor, resulting in the production of neutralizing and nonneutralizing antibodies. We found that antibody sequences from different discovery cohorts shared biochemical properties and could be retrieved across validation cohorts, confirming the stereotyped character of this naive response in coronavirus disease 2019 (COVID-19). While neutralizing antibody sequences were found independently of disease severity, in line with serological data, individual nonneutralizing antibody sequences were associated with fatal clinical courses, suggesting detrimental effects of these antibodies. We mined 200 immune repertoires from healthy individuals and 500 repertoires from patients with blood or solid cancers--all acquired prior to the pandemic--for SARS-CoV-2 antibody sequences. While the largely unmutated B cell rearrangements occurred in a substantial fraction of immune repertoires from young and healthy individuals, these sequences were less likely to be found in individuals over 60 years of age and in those with cancer. This reflects B cell repertoire restriction in aging and cancer, and may to a certain extent explain the different clinical courses of COVID-19 observed in these risk groups. Future studies will have to address if this stereotyped B cell response to SARS-CoV-2 emerging from unmutated antibody rearrangements will create long-lived memory., Introduction Severe acute respiratory syndrome coronavirus 2-specific (SARS-CoV-2-specific) B cell immunity and antibody responses are being actively investigated, since early neutralization of the virus may play a decisive role in [...]

Rickettsial diseases, scrub typhus, and Q fever are increasingly recognized as key causes of undifferentiated fevers in humans in Africa (1-6). According to a recent study in western Kenya, respiratory [...]

The transfer of T cell receptor (TCR) genes into patient T cells is a promising approach for the treatment of both viral infections and cancer. Although efficient methods exist to [...]

ABSTRACT Blastomycosis is a pulmonary disease produced in humans and animals by the thermally dimorphic fungal pathogen Blastomyces dermatitidis. In recent years, researchers and physicians have stimulated considerable interest with [...]

Immunoprecipitation followed by mass spectrometry (IP/MS) has recently emerged as a preferred method in the analysis of protein complex components and cellular protein networks. Targeting endogenous protein complexes of higher eukaryotes, particularly in large-scale efforts, has been challenging due to cellular heterogeneity, high proteome complexity, and, compared to lower organisms, lack of efficient in-locus epitope-tagging techniques. It is further complicated by variability in nonspecific identifications and cross-reactivity of primary antibodies. Still, the study of endogenous human protein networks is highly desired despite its challenges. Here we describe a streamlined IP/MS protocol for the purification and identification of extended endogenous protein complexes. We investigate the sources of nonspecific protein binding and develop semiquantitative specificity filters that are based on peptide spectral count measurements. We also outline logical constraints for the derivation of accurate complex composition from IP/MS data and demonstrate the effectiveness of this approach by presenting our analyses of different transcriptional coregulator complexes. We show consistent purification of novel components for the Integrator complex, analyze the composition of the Mediator complex solely from our data to demonstrate the wide usability of spectral counts, and deconvolute heterogeneous HDAC1/2 networks into core complex modules and several novel subcomplex interactions. antibody cross-reactivity | complex heterogeneity | protein complex | protein-protein interactions | transcriptional coregulators www.pnas.org/cgi/doi/10.1073/pnas.0912599106

Antibody repertoire diversity, potentially as high as [10.sup.11] unique molecules in a single individual, confounds characterization by conventional sequence analyses. In this study, we present a general method for assessing human antibody sequence diversity displayed on phage using massively parallel pyrosequencing, a novel application of Kabat column-labeled profile Hidden Markov Models, and translated complementarity determining region (CDR) capture-recapture analysis. Pyrosequencing of domain amplicon and RCA PCR products generated 1.5 x [10.sup.6] reads, including more than 1.9 x [10.sup.5] high quality, full-length sequences of antibody variable fragment (Fv) variable domains. Novel methods for germline and CDR classification and fine characterization of sequence diversity in the 6 CDRs are presented. Diverse germline contributions to the repertoire with random heavy and light chain pairing are observed. All germline families were found to be represented in 1.7 x [10.sup.4] sequences obtained from repeated panning of the library. While the most variable CDR (CDR-H3) presents significant length and sequence variability, we find a substantial contribution to total diversity from somatically mutated germline encoded CDRs 1 and 2. Using a capture-recapture method, the total diversity of the antibody library obtained from a human donor Immunoglobulin M (IgM) pool was determined to be at least 3.5 x [10.sup.10]. The results provide insights into the role of IgM diversification, display library construction, and productive germline usages in antibody libraries and the humoral repertoire. HMM | phage display |pyrosequencing|CDRs www.pnas.org/doi/10.1073/pnas.0909775106

The homogeneous immunosensor design described here utilizes the bivalent nature of the antibody. Antigen peptide is conjugated using flexible linkers with short complementary oligenucleotides (signaling oligonucleotides), each of which containing a fluorochrome that can form a fluorescence resonance energy transfer (FRET) donor-acceptor pair. The complementary signaling oligonucleotides are short enough to prevent their annealing on their own. Binding of the peptide-signaling oligonucleotide constructs to bivalent antibody results in a large increase in local concentration of signaling oligonucleotides causing their annealing and appearance of FRET signal. We used simple model system (antibiotin antibody) to obtain proofer-principle validation of the sensor design. We then constructed two sensors based on two peptides corresponding to the antigens of two antibodies raised against human cardiac troponin I. We demonstrated that these sensors could be used for sensitive detection of the antibody and for competition-based detection of the intact troponin I. Furthermore, we showed that these sensors could be used for detection of kinase activity targeting the antigen peptide. These simple and robust immunosensors may find applications in antibody detection (for example, in diagnosis of autoimmune or infectious disease), in protein detection (especially when speed of detection is essential), and in assays for detecting enzymatic activities involved in pest-translational modifications of proteins.

DNA aptamers carrying Pt nanoparticles were prepared by the reaction of DNA aptamers (without functionalization with biotin, thiol, or other reactive groups) with [K.sub.2][[PtCl.sub.4]] in solution at 60-90 [degrees]C. The DNA-Pt complexes possessed peroxidase enzymatic activity while retaining the specific binding ability of the aptamers. The enzymatic reaction of these complexes obeyed Michaelis-Menten kinetics. [K.sub.M] for the DNA-Pt complex was found to be on the same order as [K.sub.M] for hemin and hemin-DNA complex but 1 or 2 orders of magnitude higher than that of horseradish peroxidase. The rate of the reaction catalyzed by the DNA-Pt complex, [k.sub.cat], was found to be on the same order as that of hemin and hemin-DNA complex but 2 or 3 orders of magnitude lower than that of horseradish peroxidase. Two types of DNAzyme-linked aptamer assays (DLAAs) were developed using these complexes, which successfully detected target proteins, with the sandwich type of DLAA targeting thrombin and the competitive type of DLAA targeting anti-thrombin IgA/G/M in serum. The DNA-Pt complexes retained their peroxidase enzymatic activity even after heat treatment. DLAAs having high thermal stability were developed using these complexes, which were free of animal and plant matter because neither antibodies nor horseradish peroxidase were used in their synthesis.

The genus Henipavirus in the family Paramyxoviridae contains 2 highly lethal viruses, Hendra virus (HeV) and Nipah virus (NiV), both of which use pteropid bats as their main natural reservoir [...]

To the Editor: Filoviruses cause highly lethal hemorrhagic fever in humans and nonhuman primates, except for Reston Ebolavirus (REBOV), which causes severe hemorrhagic fever in macaques (1,2). REBOV epizootics among [...]

Our laboratory has been involved with immunodiagnostic assays for blastomycosis, a systemic dimorphilogical fungal disease of humans and animals. We have been using the ELISA tests to detect antibodies and [...]