[Thoracocentesis: wright indication and wright placement?].

The incidence of a tension pneumothorax is very low. The initial treatment of a tension pneumothorax is needle thoracocentesis. This procedure is not a routine treatment and this invasive treatment is not without risks.

Directing HIV-1 for degradation by non-target cells, using bi-specific single-chain llama antibodies.

While vaccination against HIV-1 has been so far unsuccessful, recently broadly neutralizing antibodies (bNAbs) against HIV-1 envelope glycoprotein were shown to induce long-term suppression in the absence of antiretroviral therapy in patients with antibody-sensitive viral reservoirs. The requirement of neutralizing antibodies indicates that the antibody mediated removal (clearance) of HIV-1 in itself is not efficient enough in these immune compromised patients. Here we present a novel, alternative approach that is independent of a functional immune system to clear HIV-1, by capturing the virus and redirecting it to non-target cells where it is internalized and degraded.

Non-invasive electrical cardiometry cardiac output monitoring during prehospital helicopter emergency medical care: a feasibility study.

Purpose: Introducing advanced hemodynamic monitoring might be beneficial during Helicopter Emergency Medical Service (HEMS) care. However, it should not increase the on-scene-time, it should be easy to use and should be non-invasive. The goal of this study was to investigate the feasibility of non-invasive cardiac output measurements by electrical cardiometry (EC) and the quality of the EC signal during pre-hospital care provided by our HEMS.

Nanobodies that Neutralize HIV.

Nanobodies or VHH (variable domains of heavy-chain only antibodies) are derived from camelid species such as llamas and camels. Nanobodies isolated and selected through phage display can neutralize a broad range of human immunodeficiency virus type 1 (HIV-1) strains. Nanobodies fit into canyons on the HIV envelope that may not be accessible to IgG (immunoglobulin G) containing both heavy and light chains, and they tend to have long CDR3 (complementarity-determining region 3) loops that further enhance recognition of otherwise cryptic epitopes.

Towards an ultra-rapid smartphone- connected test for infectious diseases.

The development is reported of an ultra-rapid, point-of-care diagnostic device which harnesses surface acoustic wave (SAW) biochips, to detect HIV in a finger prick of blood within 10 seconds (sample-in-result-out). The disposable quartz biochip, based on microelectronic components found in every consumer smartphone, is extremely fast because no complex labelling, amplification or wash steps are needed. A pocket-sized control box reads out the SAW signal and displays results electronically.

Unravelling the Molecular Basis of High Affinity Nanobodies against HIV p24: In Vitro Functional, Structural, and in Silico Insights.

Preventing the spread of infectious diseases remains an urgent priority worldwide, and this is driving the development of advanced nanotechnology to diagnose infections at the point of care. Herein, we report the creation of a library of novel nanobody capture ligands to detect p24, one of the earliest markers of HIV infection. We demonstrate that these nanobodies, one tenth the size of conventional antibodies, exhibit high sensitivity and broad specificity to global HIV-1 subtypes.

A novel immuno-gold labeling protocol for nanobody-based detection of HER2 in breast cancer cells using immuno-electron microscopy.

Immuno-electron microscopy is commonly performed with the use of antibodies. In the last decade the antibody fragment indicated as nanobody (VHH or single domain antibody) has found its way to different applications previously done with conventional antibodies. Nanobodies can be selected to bind with high affinity and specificity to different antigens.

Effective Inhibition of Bone Morphogenetic Protein Function by Highly Specific Llama-Derived Antibodies.

Bone morphogenetic proteins (BMP) have important but distinct roles in tissue homeostasis and disease, including carcinogenesis and tumor progression. A large number of BMP inhibitors are available to study BMP function; however, as most of these antagonists are promiscuous, evaluating specific effects of individual BMPs is not feasible. Because the oncogenic role of the different BMPs varies for each neoplasm, highly selective BMP inhibitors are required.

Camelid heavy chain only antibody fragment domain against β-site of amyloid precursor protein cleaving enzyme 1 inhibits β-secretase activity in vitro and in vivo.

Unlabelled: Accumulation and aggregation of the amyloid-β (Aβ) peptide is associated with Alzheimer's disease (AD). Aβ is generated from the amyloid precursor protein by the successive action of two membrane-associated processing enzymes: β-secretase or β-site of amyloid precursor protein cleaving enzyme 1 (BACE1) and γ-secretase. Inhibition of one or both of these enzymes prevents Aβ generation and the accompanying Aβ accumulation.

Neutralization properties of simian immunodeficiency viruses infecting chimpanzees and gorillas.

Unlabelled: Broadly cross-reactive neutralizing antibodies (bNabs) represent powerful tools to combat human immunodeficiency virus type 1 (HIV-1) infection. Here, we examined whether HIV-1-specific bNabs are capable of cross-neutralizing distantly related simian immunodeficiency viruses (SIVs) infecting central (Pan troglodytes troglodytes) (SIVcpzPtt) and eastern (Pan troglodytes schweinfurthii) (SIVcpzPts) chimpanzees (n = 11) as well as western gorillas (Gorilla gorilla gorilla) (SIVgor) (n = 1). We found that bNabs directed against the CD4 binding site (n = 10), peptidoglycans at the base of variable loop 3 (V3) (n = 5), and epitopes at the interface of surface (gp120) and membrane-bound (gp41) envelope glycoproteins (n = 5) failed to neutralize SIVcpz and SIVgor strains.

Neutralizing nanobodies targeting diverse chemokines effectively inhibit chemokine function.

Chemokine receptors and their ligands play a prominent role in immune regulation but many have also been implicated in inflammatory diseases such as multiple sclerosis, rheumatoid arthritis, allograft rejection after transplantation, and also in cancer metastasis. Most approaches to therapeutically target the chemokine system involve targeting of chemokine receptors with low molecular weight antagonists. Here we describe the selection and characterization of an unprecedented large and diverse panel of neutralizing Nanobodies (single domain camelid antibodies fragment) directed against several chemokines.

A gp41 MPER-specific llama VHH requires a hydrophobic CDR3 for neutralization but not for antigen recognition.

The membrane proximal external region (MPER) of the HIV-1 glycoprotein gp41 is targeted by the broadly neutralizing antibodies 2F5 and 4E10. To date, no immunization regimen in animals or humans has produced HIV-1 neutralizing MPER-specific antibodies. We immunized llamas with gp41-MPER proteoliposomes and selected a MPER-specific single chain antibody (VHH), 2H10, whose epitope overlaps with that of mAb 2F5.

Novel contrasting and labeling procedures for correlative microscopy of thawed cryosections.

One of the major challenges for correlative microscopy is the preparation of the sample; the protocols for transmission electron microscopy (TEM) and fluorescence microscopy (FM) often prove to be incompatible. Here, we introduce 2+Staining: an improved contrasting procedure for Tokuyasu sections that yields both excellent positive membrane contrast in the TEM and bright fluorescence of the probe labeled on the section. 2+Staining involves the contrasting of the immunolabeled sections with 1% osmium tetroxide, 2% uranyl acetate and lead citrate in sequential steps, followed by embedding in 1.

Optimizing immuno-labeling for correlative fluorescence and electron microscopy on a single specimen.

Correlative fluorescence and electron microscopy has become an indispensible tool for research in cell biology. The integrated Laser and Electron Microscope (iLEM) combines a Fluorescence Microscope (FM) and a Transmission Electron Microscope (TEM) within one set-up. This unique imaging tool allows for rapid identification of a region of interest with the FM, and subsequent high resolution TEM imaging of this area.

Kinetics of PKCε activating and inhibiting llama single chain antibodies and their effect on PKCε translocation in HeLa cells.

Dysregulation of PKCε is involved in several serious diseases such as cancer, type II diabetes and Alzheimer's disease. Therefore, specific activators and inhibitors of PKCε hold promise as future therapeutics, in addition to being useful in research into PKCε regulated pathways. We have previously described llama single chain antibodies (VHHs) that specifically activate (A10, C1 and D1) or inhibit (E6 and G8) human recombinant PKCε.

Biological and technical considerations regarding the removal of bacteriotoxins in sepsis with emphasis on toxic shock syndrome toxin 1.

Severe sepsis is characterized by rapid development of multiple organ failure associated with high mortality. Bacterial toxin release triggers a sequence of events that activates intracellular pathways to produce inflammatory mediators and nitric oxide. There have been numerous attempts to interrupt this devastating cascade by removing toxins, removing or inhibiting mediators, and by blocking receptors of mediators.

Enhancement of polymeric immunoglobulin receptor transcytosis by biparatopic VHH.

The polymeric immunoglobulin receptor (pIgR) ensures the transport of dimeric immunoglobulin A (dIgA) and pentameric immunoglobulin M (pIgM) across epithelia to the mucosal layer of for example the intestines and the lungs via transcytosis. Per day the human pIgR mediates the excretion of 2 to 5 grams of dIgA into the mucosa of luminal organs. This system could prove useful for therapies aiming at excretion of compounds into the mucosa.

Transmigration of beta amyloid specific heavy chain antibody fragments across the in vitro blood-brain barrier.

Previously selected amyloid beta recognizing heavy chain antibody fragments (VHH) affinity binders derived from the Camelid heavy chain antibody repertoire were tested for their propensity to cross the blood-brain barrier (BBB) using an established in vitro BBB co-culture system. Of all tested VHH, ni3A showed highest transmigration efficiency which is, in part, facilitated by a three amino acid substitutions in its N-terminal domain. Additional studies indicated that the mechanism of transcellular passage of ni3A is by active transport.

Llama-derived single domain antibodies to build multivalent, superpotent and broadened neutralizing anti-viral molecules.

For efficient prevention of viral infections and cross protection, simultaneous targeting of multiple viral epitopes is a powerful strategy. Llama heavy chain antibody fragments (VHH) against the trimeric envelope proteins of Respiratory Syncytial Virus (Fusion protein), Rabies virus (Glycoprotein) and H5N1 Influenza (Hemagglutinin 5) were selected from llama derived immune libraries by phage display. Neutralizing VHH recognizing different epitopes in the receptor binding sites on the spikes with affinities in the low nanomolar range were identified for all the three viruses by viral neutralization assays.

VIS2FIX: a high-speed fixation method for immuno-electron microscopy.

Immuno-transmission electron microscopy (TEM) is the technique of choice for high-resolution localization of proteins in fixed specimen. Here we introduce 2 novel methods for the fixation of sections from cryo-immobilized samples that result in excellent ultrastructural preservation. These high-speed fixation techniques, both called VIS2FIX, allow for a reduction in sample preparation time from at least 1 week to only 8 h.

The development of activating and inhibiting camelid VHH domains against human protein kinase C epsilon.

The 10 isozymes of the protein kinase C (PKC) family can have different roles on the same biological process, making isozyme specific analysis of function crucial. Currently, only few pharmacological compounds with moderate isozyme specific effects exist thus hampering research into individual PKC isozymes. The antigen binding regions of camelid single chain antibodies (VHHs) could provide a solution for obtaining PKC isozyme specific modulators.

Llama heavy-chain antibody fragments efficiently remove toxic shock syndrome toxin 1 from plasma in vitro but not in experimental porcine septic shock.

Staphylococcus aureus produces the superantigen toxic shock syndrome toxin 1 (TSST-1). When the bacterium invades the human circulation, this toxin can induce life-threatening gram-positive sepsis. Current sepsis treatment does not remove bacterial toxins.

Differential recognition of vascular and parenchymal beta amyloid deposition.

By phage display, llama-derived heavy chain antibody fragments were selected from non-immune and immune libraries and tested for their affinity and specificity for beta amyloid by phage-ELISA, immunohistochemistry and surface plasmon resonance. We identified eight distinct heavy chain antibody fragments specific for beta amyloid. While three of them recognized vascular and parenchymal beta amyloid deposits, the remaining five heavy chain antibody fragments recognized vascular beta amyloid specifically, failing to bind to parenchymal beta amyloid.

Reverse proteomic antibody screening identifies anti adhesive VHH targeting VLA-3.

Therapeutic approaches aimed at targeting tumor surface markers using monoclonal antibodies provide a powerful strategy in cancer treatment. Here we report selection of single variable domains (VHH) of llama heavy chain antibodies, using a VHH-phage-display library. A reverse proteomic approach was used to identify the cognate proteins recognized by enriched VHH on HeLa cells.