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Toxins May 2024Scorpion envenomation poses a global public health issue, with an estimated 1,500,000 cases worldwide annually resulting in 2600 deaths. North Africa, particularly...
Scorpion envenomation poses a global public health issue, with an estimated 1,500,000 cases worldwide annually resulting in 2600 deaths. North Africa, particularly Morocco, experiences severe envenomations, mainly attributed to and in Morocco, and and in Algeria and Tunisia, with case numbers often underestimated. Current treatment relies mainly on symptomatic approaches, except in Morocco, where management is limited to symptomatic treatment due to controversies regarding specific treatment. In Morocco, between 30,000 and 50,000 scorpion envenomation cases are reported annually, leading to hundreds of deaths, mainly among children. Controversies among clinicians persist regarding the appropriate course of action, often limiting treatments to symptomatic measures. The absence of a specific antivenom for the venoms of the most lethal scorpions further exacerbates the situation. This study aims to address this gap by developing a monovalent antivenom against the endemic and most dangerous scorpion, . The antivenom was produced by immunizing albino rabbits with a mixture of venom collected from high-risk areas in Morocco. Immunizations were performed by subcutaneous injections at multiple sites near the lymphatic system, following an immunization schedule. Production control of neutralizing antibody titers was conducted through immunodiffusion. Once a sufficient antibody titer was achieved, blood collection was performed, and the recovered plasma underwent affinity chromatography. The efficacy of purified IgG was evaluated by determining the ED in mice, complemented by histological and immunohistochemical studies on its ability to neutralize venom-induced tissue alterations and the neutralization of toxins bound to receptors in the studied organs. The monovalent antivenom demonstrated specificity against venom and effective cross-protection against the venom of the scorpions and , highly implicated in lethal envenomations in the Maghreb. This study shows that the developed monovalent antivenom exhibits notable efficacy against local scorpions and a surprising ability to neutralize the most lethal envenomations in North Africa. These results pave the way for a new, more specific, and promising therapeutic approach to countering severe scorpion envenomations, especially in Morocco, where specific treatment is lacking.
Topics: Animals; Humans; Africa, Northern; Antivenins; Morocco; Scorpion Stings; Scorpion Venoms; Scorpions
PubMed: 38787066
DOI: 10.3390/toxins16050214 -
Journal of Fungi (Basel, Switzerland) Apr 2024Coccidiomycosis is a potentially life-threatening fungal infection endemic to certain regions of Argentina. The infection is caused by spp. and is primarily diagnosed...
Coccidiomycosis is a potentially life-threatening fungal infection endemic to certain regions of Argentina. The infection is caused by spp. and is primarily diagnosed by antibody (Ab) detection. Access to rapid, highly accurate diagnostic testing is critical to ensure prompt antifungal therapy. The sōna Ab Lateral Flow Assay (LFA) performs faster and requires less laboratory infrastructure and equipment compared with other Ab detection assays, potentially providing a substantial improvement for rapid case screening in coccidioidomycosis-endemic regions; however, validation of this test is needed. Thus, we aimed to evaluate the analytical performance of the sōna Ab (LFA) and compare agreement with anti- Ab detection assays. A total of 103 human sera specimens were tested, including 25 specimens from patients with coccidioidomycosis and 78 from patients without coccidioidomycosis. The sōna Ab Lateral Flow Assay (LFA) was performed with a sensitivity of 88%, and specificity and accuracy of 87%. Furthermore, the Ab LFA had good agreement with other anti- Ab detection assays. Our findings suggest the sōna Ab LFA has satisfactory performance and may be useful for diagnosing coccidioidomycosis in endemic regions.
PubMed: 38786677
DOI: 10.3390/jof10050322 -
Cells May 2024Monocytes, as well as downstream macrophages and dendritic cells, are essential players in the immune system, fulfilling key roles in homeostasis as well as in...
Monocytes, as well as downstream macrophages and dendritic cells, are essential players in the immune system, fulfilling key roles in homeostasis as well as in inflammatory conditions. Conventionally, driven by studies on reporter models, mouse monocytes are categorized into a classical and a non-classical subset based on their inversely correlated surface expression of Ly6C/CCR2 and CX3CR1. Here, we aimed to challenge this concept by antibody staining and reporter mouse models. Therefore, we took advantage of and reporter mice, in which the respective gene was replaced by a fluorescent reporter protein gene. We analyzed the expression of CX3CR1 and CCR2 by flow cytometry using several validated fluorochrome-coupled antibodies and compared them with the reporter gene signal in these reporter mouse strains. Although we were able to validate the specificity of the fluorochrome-coupled flow cytometry antibodies, mouse Ly6C classical and Ly6C non-classical monocytes showed no differences in CX3CR1 expression levels in the peripheral blood and spleen when stained with these antibodies. On the contrary, in reporter mice, we were able to reproduce the inverse correlation of the CX3CR1 reporter gene signal and Ly6C surface expression. Furthermore, differential CCR2 surface expression correlating with the expression of Ly6C was observed by antibody staining, but not in reporter mice. In conclusion, our data suggest that phenotyping strategies for mouse monocyte subsets should be carefully selected. In accordance with the literature, the suitability of CX3CR1 antibody staining is limited, whereas for CCR2, caution should be applied when using reporter mice.
Topics: Animals; Receptors, CCR2; Monocytes; CX3C Chemokine Receptor 1; Mice; Flow Cytometry; Antibodies; Genes, Reporter; Phenotype; Mice, Inbred C57BL; Mice, Transgenic; Green Fluorescent Proteins; Antigens, Ly
PubMed: 38786041
DOI: 10.3390/cells13100819 -
Biosensors May 2024Loop-mediated isothermal amplification (LAMP) technology is extensively utilized for the detection of infectious diseases owing to its rapid processing and high...
Loop-mediated isothermal amplification (LAMP) technology is extensively utilized for the detection of infectious diseases owing to its rapid processing and high sensitivity. Nevertheless, conventional LAMP signaling methods frequently suffer from a lack of sequence specificity. This study integrates a triplex-forming oligonucleotide (TFO) probe into the LAMP process to enhance sequence specificity. This TFO-LAMP technique was applied for the detection of Group B (GBS). The TFO probe is designed to recognize a specific DNA sequence, termed the TFO targeting sequence (TTS), within the amplified product, facilitating detection via fluorescent instrumentation or lateral flow biosensors. A screening method was developed to identify TFO sequences with high affinity to integrate TFO into LAMP, subsequently incorporating a selected TTS into an LAMP primer. In the TFO-LAMP assay, a FAM-labeled TFO is added to target the TTS. This TFO can be captured by an anti-FAM antibody on lateral flow test strips, thus creating a nucleic acid testing biosensor. The efficacy of the TFO-LAMP assay was confirmed through experiments with specimens spiked with varying concentrations of GBS, demonstrating 85% sensitivity at 300 copies and 100% sensitivity at 30,000 copies. In conclusion, this study has successfully developed a TFO-LAMP technology that offers applicability in lateral flow biosensors and potentially other biosensor platforms.
Topics: Biosensing Techniques; Nucleic Acid Amplification Techniques; Oligonucleotides; Streptococcus; Humans; DNA, Bacterial; Molecular Diagnostic Techniques
PubMed: 38785731
DOI: 10.3390/bios14050257 -
Biosensors May 2024Polluted air and the presence of numerous airborne pathogens affect our daily lives. The sensitive and fast detection of pollutants and pathogens is crucial for...
Polluted air and the presence of numerous airborne pathogens affect our daily lives. The sensitive and fast detection of pollutants and pathogens is crucial for environmental monitoring and effective medical diagnostics. Compared to conventional detection methods (PCR, ELISA, metabolic tests, etc.), biosensors bring a very attractive possibility to detect chemicals and organic particles with the mentioned reliability and sensitivity in real time. Moreover, by integrating nanomaterials into the biosensor structure, it is possible to increase the sensitivity and specificity of the device significantly. However, air quality monitoring could be more problematic even with such devices. The greatest challenge with conservative and sensing methods for detecting organic matter such as bacteria is the need to use liquid samples, which slows down the detection procedure and makes it more difficult. In this work, we present the development of a polyacrylonitrile nanofiber bioreceptor functionalized with antibodies against bacterial antigens for the specific interception of bacterial cells directly from the air. We tested the presented novel nanofiber bioreceptor using a unique air filtration system we had previously created. The prepared antibody-functionalized nanofiber membranes for air filtration and pathogen detection (with model organisms and ) show a statistically significant increase in bacterial interception compared to unmodified nanofibers. Creating such a bioreceptor could lead to the development of an inexpensive, fast, sensitive, and incredibly selective bionanosensor for detecting bacterial polluted air in commercial premises or medical facilities.
Topics: Nanofibers; Staphylococcus aureus; Biosensing Techniques; Escherichia coli; Environmental Monitoring; Air Microbiology; Acrylic Resins
PubMed: 38785708
DOI: 10.3390/bios14050234 -
Microsystems & Nanoengineering 2024The development of artificial intelligence-enabled medical health care has created both opportunities and challenges for next-generation biosensor technology. Proteins... (Review)
Review
The development of artificial intelligence-enabled medical health care has created both opportunities and challenges for next-generation biosensor technology. Proteins are extensively used as biological macromolecular markers in disease diagnosis and the analysis of therapeutic effects. Electrochemical protein biosensors have achieved desirable specificity by using the specific antibody-antigen binding principle in immunology. However, the active centers of protein biomarkers are surrounded by a peptide matrix, which hinders charge transfer and results in insufficient sensor sensitivity. Therefore, electrode-modified materials and transducer devices have been designed to increase the sensitivity and improve the practical application prospects of electrochemical protein sensors. In this review, we summarize recent reports of electrochemical biosensors for protein biomarker detection. We highlight the latest research on electrochemical protein biosensors for the detection of cancer, viral infectious diseases, inflammation, and other diseases. The corresponding sensitive materials, transducer structures, and detection principles associated with such biosensors are also addressed generally. Finally, we present an outlook on the use of electrochemical protein biosensors for disease marker detection for the next few years.
PubMed: 38784375
DOI: 10.1038/s41378-024-00700-w -
Sheng Wu Gong Cheng Xue Bao = Chinese... May 2024The antibodies to the microtubule-associated protein tau play a role in basic and clinical studies of Alzheimer's disease (AD) and other tauopathies. With the...
The antibodies to the microtubule-associated protein tau play a role in basic and clinical studies of Alzheimer's disease (AD) and other tauopathies. With the recombinant human tau441 as the immunogen, the hybridoma cell strains secreting the anti-human tau N-terminal domain (NTD-tau) monoclonal antibodies were generated by cell fusion and screened by limiting dilution. The purified monoclonal antibodies were obtained by inducing the mouse ascites and affinity chromatography. The sensitivity and specificity of the monoclonal antibodies were examined by indirect ELISA and Western blotting, respectively. A double antibody sandwich ELISA method for detecting human tau protein was established and optimized. The results showed that the positive cloning rate of hybridoma cells was 83.6%. A stable cell line producing ZD8F7 antibodies was established, and the antibody titer in the supernatant of the cell line was 1:16 000. The antibody titer in the ascitic fluid was higher than 1:256 000; and the titer of purified ZD8F7 monoclonal antibodies was higher than 1:128 000. The epitope analysis showed that the ZD8F7 antibody recognized tau21-37 amino acid in the N-terminal domain. The Western blotting results showed that the ZD8F7 antibody recognized the recombinant human tau protein of 50-70 kDa and the human tau protein of 50 kDa in the brain tissue of transgenic AD model mice (APP/PS1/tau). With ZD8F7 as a capture antibody, a quantitative detection method for human tau protein was established, which showed a linear range of 7.8-500.0 pg/mL and could identify human tau protein in the brain tissue of AD transgenic mice and human plasma but not recognize the mouse tau protein. In conclusion, the human NTD-tau-specific monoclonal antibody and the double antibody sandwich ELISA method established in this study are highly sensitive and can serve as a powerful tool for the detection of tau protein in neurodegenerative diseases.
Topics: tau Proteins; Animals; Antibodies, Monoclonal; Humans; Mice; Alzheimer Disease; Enzyme-Linked Immunosorbent Assay; Recombinant Proteins; Hybridomas; Mice, Inbred BALB C; Antibody Specificity; Protein Domains; Epitopes
PubMed: 38783817
DOI: 10.13345/j.cjb.230655 -
Journal of Microbiology and... Jun 2024Protein-specific antibodies are essential for various aspects of protein research, including detection, purification, and characterization. When specific antibodies are...
Protein-specific antibodies are essential for various aspects of protein research, including detection, purification, and characterization. When specific antibodies are unavailable, protein tagging is a useful alternative. Small epitope tags, typically less than 10 amino acids, are widely used in protein research due to the simple modification through PCR and reduced impact on the target protein's function compared to larger tags. The 2B8 epitope tag (RDPLPFFPP), reported by us in a previous study, has high specificity and sensitivity to the corresponding antibody. However, when attached to the C-terminus of the target protein in immunoprecipitation experiments, we observed a decrease in detection signal with reduced immunity and low protein recovery. This phenomenon was not unique to 2B8 and was also observed with the commercially available Myc tag. Our study revealed that C-terminal tagging of small epitope tags requires the addition of more than one extra amino acid to enhance (restore) antibody immunities. Moreover, among the amino acids we tested, serine was the best for the 2B8 tag. Our findings demonstrated that the interaction between a small epitope and a corresponding paratope of an antibody requires an extra amino acid at the C-terminus of the epitope. This result is important for researchers planning studies on target proteins using small epitope tags.
Topics: Epitopes; Amino Acids; Animals; Antibodies; Mice; Immunoprecipitation; Antibody Formation; Recombinant Fusion Proteins
PubMed: 38783697
DOI: 10.4014/jmb.2401.01036 -
Journal of Clinical Virology : the... Aug 2024HDV antibody testing is recommended for universal screening and as the first line in an HDV double reflex testing strategy for effectively identifying patients with...
BACKGROUND
HDV antibody testing is recommended for universal screening and as the first line in an HDV double reflex testing strategy for effectively identifying patients with active infection for therapeutic treatments.
OBJECTIVE
The aim of this study is to evaluate the performance of a newly developed ARCHITECT HDV Total Ig (ARCHITECT HDV Ig) prototype assay.
STUDY DESIGN
Performance characteristics were determined for the ARCHITECT HDV Ig and a reference test, LIAISON XL Anti-HDV using a well-characterized specimen panel, comprising HDV RNA positive (n = 62) and negative (n = 70) samples, and healthy US blood donors.
RESULTS
Healthy US blood donors (n=200) showed 99.5% (199/200, 95%CI=97.65-99.98) specificity with ARCHITECT HDV Ig and 98.5 % (197/200, 95 %CI = 96.10-99.64) with LIAISON Anti-HDV. Among known HDV RNA positive samples, ARCHITECT HDV Ig detected 59/62 demonstrating 95.2 % sensitivity while LIAISON Anti-HDV sensitivity was 90.3 % (56/62). Among 101 HBV positive samples, 70 were reactive in the ARCHITECT test, 59 of which tested positive for HDV RNA for a positive predictive value (PPV) for the presence of HDV RNA was 84.3 %. For LIAISON Anti-HDV, 79 specimens were reactive and 56 contained HDV RNA: PPV for HDV RNA was 70.9 %. Among 70 HDV RNA negative samples, 39 were HBV positive. ARCHITECT HDV Ig negative predictive value (NPV) was 71.8 % and LIAISON Anti-HDV NPV was 41 % for the HBV positive group, respectively.
CONCLUSION
When compared to the LIASON Anti-HDV test, the ARCHITECT HDV Ig assay demonstrated enhanced sensitivity and specificity and better NPV and PPV values for HDV RNA status. The ARCHITECT HDV Ig assay represents a promising tool for universal screening of all HBsAg-positive persons.
Topics: Humans; Sensitivity and Specificity; Hepatitis D; Hepatitis Delta Virus; Hepatitis Antibodies; High-Throughput Screening Assays; Serologic Tests; Automation, Laboratory; Blood Donors
PubMed: 38781633
DOI: 10.1016/j.jcv.2024.105689 -
Proceedings of the National Academy of... May 2024The HapImmune platform exploits covalent inhibitors as haptens for creating major histocompatibility complex (MHC)-presented tumor-specific neoantigens by design,...
The HapImmune platform exploits covalent inhibitors as haptens for creating major histocompatibility complex (MHC)-presented tumor-specific neoantigens by design, combining targeted therapies with immunotherapy for the treatment of drug-resistant cancers. A HapImmune antibody, R023, recognizes multiple sotorasib-conjugated KRAS(G12C) peptides presented by different human leukocyte antigens (HLAs). This high specificity to sotorasib, coupled with broad HLA-binding capability, enables such antibodies, when reformatted as T cell engagers, to potently and selectively kill sotorasib-resistant KRAS(G12C) cancer cells expressing different HLAs upon sotorasib treatment. The loosening of HLA restriction could increase the patient population that can benefit from this therapeutic approach. To understand the molecular basis for its unconventional binding capability, we used single-particle cryogenic electron microscopy to determine the structures of R023 bound to multiple sotorasib-peptide conjugates presented by different HLAs. R023 forms a pocket for sotorasib between the V and V domains, binds HLAs in an unconventional, angled way, with V making most contacts with them, and makes few contacts with the peptide moieties. This binding mode enables the antibody to accommodate different hapten-peptide conjugates and to adjust its conformation to different HLAs presenting hapten-peptides. Deep mutational scanning validated the structures and revealed distinct levels of mutation tolerance by sotorasib- and HLA-binding residues. Together, our structural information and sequence landscape analysis reveal key features for achieving MHC-restricted recognition of multiple hapten-peptide antigens, which will inform the development of next-generation therapeutic antibodies.
Topics: Humans; Peptides; HLA Antigens; Major Histocompatibility Complex; Haptens; Protein Binding; Cryoelectron Microscopy
PubMed: 38781214
DOI: 10.1073/pnas.2319029121