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MALDI-TOF MS is well-recognized for microbial identification and widely used in research and clinical fields due to its specificity, speed of analysis, and low cost of consumables. However, the classification or identification accuracy is poor for E. coli and Shigella. In addition, FTIR is a promising tool for bacterial typing. In this study, 14 strains of E. coli and 9 strains of Shigella were typed by both MALDI-TOF MS and FTIR techniques. Alternatively, a data fusion strategy using these two approaches was attempted to achieve better typing accuracy. Hierarchical clustering analysis (HCA) revealed that the typing accuracies for selected E. coli and Shigella from blood agar were 65.2%, 78.3%, and 100% for MALDI-TOF MS, FTIR, and FTIR combined with MALDI-TOF MS, respectively. Based on these results, a library constructed of the MS-IR fusion data was employed to identify individual bacteria at the strain level, and the library-based typing accuracies for 207 test spectra of sample strains from three different culture media yielded 97.6% accuracy for distinguishing between E. coli and Shigella at genus level. For species level and strain level, it yielded 95.2% and 92.3% typing accuracy, respectively. These results indicate that FTIR is a better technique for typing selected E. coli and Shigella than MALDI-TOF MS, and the accuracy is increased by combining the data from MALDI-TOF MS and FTIR spectroscopy. FTIR could be used to supplement MALDI-TOF MS for the identification and typing of taxonomic microorganisms.

Tumor-derived exosomes, which contain RNA, DNA, and proteins, are a potentially rich non-invasive source of biomarkers. However, no efficient isolation or detection methods are yet available. Here, we developed a microfluidic Raman biochip designed to isolate and analyze exosomes in situ. Anti-CD63 magnetic nanoparticles were used to enrich exosomes through mixing channels of a staggered triangular pillar array. EpCAM-functionalized Raman-active polymeric nanomaterials (Raman beads) allow rapid analysis of exosome samples within 1 h, with a quantitative signal at 2230 cm-1. The limit of detection of this biochip approaches 1.6 × 102 particles per mL with 20 μL samples. The newly developed biochip assay was successfully applied in the determination of exosomes from clinical serum samples. Thus, this novel device may have potential as a clinical exosome analysis tool for prostate cancer.

Tumor exosomes that inherit specific molecules from their parent cells are emerging as ideal biomarkers in cancer diagnostics. Most currently available exosome isolation and detection methods are time-consuming and non-specific; thus, rapid and specific exosome detection methods are needed both clinically and in research. Here, a dual-functional platform is reported composed of reversible conjunction and “off-on” signal responses. Fe3O4@SiO2@TiO2 particles with high affinity were applied to capture exosomes, and model exosomes could be isolated from solution within 20 min with a capture efficiency of 91.5%. An “on-off” fluorescence response PSMA aptasensor was constructed with improved selectivity to detect tumor exosomes by recording the fluorescence intensity with λex/em = 557/580 nm. The standard curve for detecting tumor exosomes with the aptasensor was calculated as y = 371.7x + 66.17, ranging from 0.05 to 1 × 104 particles/μL, with R2 = 0.9737, and a detection limit of 5 × 102 particles/μL in solution. This method was successfully applied to clinical samples, and the results showed better performance in distinguishing prostate cancer patients and healthy samples than the traditional nanoparticle-tracking analysis (NTA) method. This rapid and accurate detection method for prostate cancer may aid in rapid clinical diagnosis.

Posttraumatic infections can occur in orthopedic trauma patients, especially in open fractures. Rapid and accurate identification of pathogens in orthopedic trauma is important for clinical diagnosis and antimicrobial treatment. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been successfully used for first-line identification of pathogens grown on culture plates. However, for direct analysis of liquid clinical specimens, pre-purification of the sample is necessary. Herein, we investigated the feasibility of coupling Fc-MBL@Fe3O4 enrichment with MALDI-TOF MS profiling in the identification of pathogens in liquid-cultured samples. This method is successfully used for the identification of pathogens in a patient with an open-leg fracture obtained at sea. Pathogens were enriched by Fc-MBL@Fe3O4 from briefly pre-cultured liquid media and identified by MALDI-TOF MS. We identified an opportunistic pathogen, Vibrio alginolyticus, which is uncommon in clinical orthopedic trauma infection but exists widely in the sea. Therefore, combining Fc-MBL@Fe3O4 enrichment and MALDI-TOF MS profiling has great potential for direct identification of microbes in clinical samples.

Enantiomeric drugs are widely used and play important roles in pharmaceuticals. Ion mobility spectrometry coupled with MS technology provides a unique method for distinguishing the enantiomeric drugs, enantiomeric identification and quantitation in the gas phase. In this study, enantiomeric molecules of ibuprofen and flurbiprofen were clearly recognized by forming host-guest complex ions using trapped ion mobility time-of-flight MS. Ternary complex ions can be produced easily by electrospray ionization of the mixed solutions of ibuprofen, cyclodextrins, and CaCl2 , LiCl, or NaCl, as well as flurbiprofen, cyclodextrins, and CaCl2 . The relative contents of different chiral ibuprofens in a mixed solution were also quantitatively measured. This new method is a simple, effective, and a convenient enantioselective analysis method. This article is protected by copyright. All rights reserved.

Bacterial infections are the key cause of morbidity and mortality worldwide. Matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS)-based bacterial identification has been widely accepted in the clinic. Functional material, such as rabbit immunoglobulin G-modified Fe3O4 (IgG@Fe3O4) and fragment crystallizable mannose binding lectin-modified Fe3O4 (FcMBL@Fe3O4), is used to capture bacteria from biological samples for MALDI-TOF MS identification, and the bacteria MS signals are usually obtained by directly smearing enriched bacteria on a MALDI target with MALDI matrix solution. However, the accuracy of identification based on MALDI-TOF MS may be affected by the presence of functional molecules, especially proteins, resulting in errors in the comparison with the standard bacterial spectra in the database. Moreover, the long-term presence of the magnetic beads on the MALDI-TOF target may reduce the instrument service life. In this study, we constructed FcMBL@Fe3O4 and used it to capture bacteria from both aqueous solution and bovine blood, and the bacterial identification accuracy based on different target preparation methods was compared. In the presence of Ca2+, the similarity scores for bacteria identified with FcMBL@Fe3O4 were ~88% and ~82% for Staphylococcus. aureus and Escherichia coli, respectively. In the presence of ethylenediaminetetraacetic acid (EDTA), bacteria separate from FcMBL@Fe3O4, resulting in similarity scores of ~96% and ~92% for S. aureus and E. coli, respectively. These results indicate that the functional proteins on the surface of nanoparticles affect the accuracy of identification accuracy based on the MALDI-TOF MS database. Thus, the release of bacteria from the functional material could increase the identification accuracy and be beneficial for maintaining the instrument.

The indiscriminate application of antibiotics has led to the spread of antibiotic-resistant bacteria, such as methicillin-resistant Staphylococcus aureus (MRSA) and Klebsiella pneumoniae carbapenemase 2-expressing Klebsiella pneumoniae (KPC-2 KP), resulting in a global threat to public health. A method for specific, sensitive, and rapid detection of such bacteria is essential for clinical diagnosis and treatment. In this study, target bacteria were selectively captured using aptamer-coated magnetic beads (Apt-MBs), followed by recognition and detection by a broad-spectrum fluorescent probe, fragment crystallizable mannose binding lectin-coated gold nanoclusters (FcMBL@AuNCs). As a proof of concept study, a Gram-positive bacterium (i.e., MRSA) and Gram-negative bacterium (i.e., KPC-2 KP) were selected as models and detected by the proposed strategy. The determination of MRSA and KPC-2 KP was in the range of 20–108 CFU/mL with a detection limit (DL) of ~20 CFU/mL. Furthermore, the method shows great specificity for target bacteria, and detection procedures could be completed in 15 min. With these priorities, this platform has been applied successfully to the detection of MRSA and KPC-2 KP in human urine, saliva, cerebrospinal fluid, and bovine blood. Utilizing different aptamers, this strategy could also be used to monitor other target bacteria with high specificity and sensitivity.

A surface-enhanced Raman scattering (SERS)-based immunocapture nanoprobe is described for the detection of pathogenic bacteria. The probe uses boronic acid-functionalized polydopamine-coated Au@Ag nanoparticles as an advanced SERS nanotag. Modified magnetic IgG@Fe3O4 nanoparticles are used for magnetic separation. Au@Ag@PDA nanoparticles, where PDA stands for polydopamine, were functionalized with boronic acid to bind to pathogenic bacteria and induce signal amplification. The Raman signal is amplified 108 times when the SERS tag binds the surface of bacteria. The SERS spectra exhibit fingerprint-like patterns that enable bacterial classification. The results of principal component analysis (PCA) and hierarchical cluster analysis (HCA) of the spectral regions were compared. The bacterial surface protein and glycan signals (1300–1450 cm−1) were the best regions for bacterial classification. Staphylococcus aureus, Escherichia coli, Shigella dysenteriae, Pseudomonas aeruginosa, and Klebsiella pneumonia were successfully classified by this method. The lowest detection limit was 10 colonies/mL (CFU·mL). The assay can be completed within 30 min. Conceivably, this method may be extended to the quantitative detection or classification of bacteria under various other conditions.

Sactionine-containing antibiotics (sactibiotics) are a growing class of peptide antibiotics belonging to the ribosomally synthesized and post-translationally modified peptide (RiPP) superfamily. We report the characterization of thuricin Z, a novel sactibiotic from Bacillus thuringiensis. Unusually, the biosynthesis of thuricin Z involves two radical S-adenosylmethionine (SAM) enzymes, ThzC and ThzD. Although ThzC and ThzD are highly divergent from each other, these two enzymes produced the same sactionine ring in the precursor peptide ThzA in vitro. Thuricin Z exhibits narrow-spectrum antibacterial activity against Bacillus cereus. A series of analyses, including confocal laser scanning microscopy, ultrathin-sectioning transmission electron microscopy, scanning electron microscopy, and large-unilamellar-vesicle-based fluorescence analysis, suggested that thuricin Z binds to the bacterial cell membrane and leads to membrane permeabilization.

Nanoparticles (NP) in biological fluids almost invariably become coated with proteins to form protein coronas. It is the NP–protein corona rather than the bare nanoparticle that determines the nanoparticle's bio-behavior. Here, ultrasmall gold nanoparticles (AuNPs) coated by a human serum albumin (HSA) corona were studied by Fourier transform infrared spectroscopy, denature experiments, fluorescence quenching. Moreover, the intracellular fate of AuNPs and the AuNP–HSA corona has also been investigated. The results show that HSA corona undergo a conformational transition (partial β-sheet changed to α-helicity) when they adsorb on AuNPs, which lead to an enhanced thermal stability. Importantly, we observed that the conformation-transited protein corona–AuNP complex could induce cell apoptosis. Meanwhile, for the first time, the conformation-transited HSA on the AuNPs surface are shown to disrupt living cell membranes. The results obtained here not only provide the detailed conformational behavior of HSA molecules on nanoparticles, but also reveal the structure–function relationship of protein corona, which is of utmost importance in the safe application of nanoscale objects in living organisms.

The serum MALDI-TOF MS spectrum includes signals for serum proteins and peptides between 1000 and 12,000 Da in size, presenting a fingerprint-like pattern. However, whole serum MALDI-TOF MS signals are complex and prejudiced for data analysis. Pre-treatment with specific nanomaterials can simplify the mass spectrum while retaining the characteristics of the fingerprint pattern. In the present study, we used hydrophilic interaction chromatography nanoparticles (HICNPs) to enrich proteins and peptides in serum from a large number prostate cancer samples and controls. After pre-treatment with HICNPs, the serum MALDI-TOF MS signals for samples were simpler, with more analysable fingerprint-like patterns. Principal component analysis and partial least squares discriminant analysis of the samples demonstrated a significant difference in the MALDI-TOF signals between prostate cancer and controls, with an analytical accuracy of 77%, approaching that of methods based on prostate-specific antigen. Due to the low cost and high flux, MALDI-TOF MS fingerprinting can be used in large-scale evaluation of various cancers, including prostate cancer.

One of the main diseases contributing to human death are malignant tumors. Phototherapy is a promising approach for cancer therapy, and functional nanoparticles with targeting ligands are commonly used to improve the therapeutic efficiency. However, recent studies have shown that nanoparticles in contact with a biological fluid can rapidly form a "protein corona" on their surface, which will remarkably decrease the targeting ability. Here, we describe the preparation of hybrid nanomaterials with Bi2S3 nanorods as the core, and fluorescein-isothiocyanate and folic acid-modified human serum albumin (HSA-FITC-FA) as the shell. By using fluorescent binding label (FITC) and imaging techniques, we discovered the image of the cell lysosomes, indicating that the photothermal therapy agent was predominantly targeted to and accumulated in lysosomes. Combined with photothermal therapy agent (Bi2S3 nanorods) and targeting ligand (FA), the obtained product shows enhanced photothermal therapy under near-infrared region laser irradiation. Additionally, SDS-PAGE shows that the modified HSA shell could remarkably reduce the reabsorption of protein corona from blood serum, minimized the adverse effect of protein corona on targetability. Taken together, the results indicate that our strategy has the potential for preparing efficient photothermal nanomaterials with image-guided subcellular organelle-targeting cancer cell ablation ability.

A method is described for fast identification of bacteria by combining (a) the enrichment of bacterial cells by using magnetite (Fe3O4) magnetic beads modified with human IgG (IgG@Fe3O4) and (b) MALDI-TOF MS analysis. IgG has affinity to protein A, protein G, protein L and glycans on the surface of bacterial cells, and IgG@Fe3O4. It therefore is applicable to the preconcentration of a range of bacterial species. The feasibility of the method has been demonstrated by collecting six species of pathogenic bacteria (Gram-positives: Staphylococcus aureus and Kocuria rosea; Gram-negatives: Klebsiella pneumoniae, Klebsiella oxytoca, Enterobacter cloacae and Pseudomonas aeruginosa). Bacteria with concentrations as low as 10 CFU·mL−1 in spiked water samples were extracted by this sorbent with recovery rates of >50%. After enrichment, bacteria on the IgG@Fe3O4 sorbent were further identified by MALDI-TOF MS. Bacteria in concentrations as low as 105 CFU in 100 μL of human whole blood can be identified by the method. Compared to other blood culture based tests, the culture time is shortened by 40% (from ~10 h to ~6 h), and the plate culture procedure (overnight) is avoided. After short blood culture, the enrichment and identification can be finished in one hour. The IgG@Fe3O4 is of practical value in clinical diagnosis and may be combined with other identification methods, e.g. PCR, Raman spectroscopy, infrared spectroscopy, etc.

Nanoparticle (NP) surfaces are modified immediately by the adsorption of proteins when exposed to human blood, leading to the formation of a protein corona. The adsorption of serum proteins is the key process for exploring the bioapplication and biosafety of NPs. In this study, NP–protein binding affinity (Ka) was investigated. Some serum proteins, such as human serum albumin (HSA), trypsin (TRP), hemoglobin (Hb), myoglobin (MB), immunoglobulin G (IgG), carbonic anhydrase (CA), fibrinogen (FIB), chymotrypsin and r-globulin, were used with gold nanoparticles (AuNPs) to address binding affinity according to isothermal titration calorimetry (ITC) combined with dynamic light scattering (DLS) and fluorescence quenching. The NP protein binding affinities determined by the two methods were in agreement, and depended on the protein properties and size of the NPs. The two methods are convenient, and the results are highly comparable. These methods can be extended to determine the binding affinity of NP protein interactions. The adsorption of proteins upon the AuNP surface is a complex process and depends on several factors, but the binding affinities are higher for proteins with more cysteine residues located on the surface.

The integration of photothermal therapy (PTT) with photodynamic therapy (PDT) has great potential to revolutionize conventional therapeutic approaches for cancers. However, most of the currently developed nanoplatforms for combinatorial phototherapy require the participation of different components or activation of separate lasers. Therefore, a desirable photoactive agent with intrinsic single laser-activated combinatorial phototherapy is still urgently needed for more efficient treatment. Here, we demonstrate a high-temperature organic synthesis, ligand exchange strategy to obtain ultrasmall, water-dispersible, and stable polylysine-bismuth sulfide quantum. dots (PLL-Bi2S3 QDs). In addition to its strong near-infrared (NIR) absorption, the ultrasmall size and sulfide-enriched surface of the obtained Bi2S3 QDs offer abundant photoelectrons with high reducing ability, promoting the rapid generation of reactive oxygen species (ROS). Therefore, excellent photothermal conversion and fast ROS generation ability can be concurrently realized upon 808 nm NIR laser irradiation. As proof of concept, the combined photothermal/photodynamic activity in single PLL-Bi2S3 QDs was demonstrated both in vitro and in vivo, confirming that the phototherapeutic efficacy is dramatically greater than that of larger Bi2S3 nanorods. In addition, toxicity screening showed that PLL-Bi2S3 QDs possess excellent biocompatibility. Thus, the prepared PLL-Bi2S3 QDs are expected to serve as a photoactive agent for efficient cancer treatment.

Targeting peptide-modified magnetic graphene-based mesoporous silica (MGMSPI) are synthesized, characterized, and developed as a multifunctional theranostic platform. This system exhibits many merits, such as biocompatibility, high near-infrared photothermal heating, facile magnetic separation, large T2 relaxation rates (r2), and a high doxorubicin (DOX) loading capacity. In vitro and in vivo results demonstrate that DOX-loaded MGMSPI (MGMSPID) can integrate magnetic resonance imaging, dual-targeting recognition (magnetic targeting and receptor-mediated active targeting), and chemo-photothermal therapy into a single system for a visualized-synergistic therapy of glioma. In addition, it is observed that the MGMSPID system has heat-stimulated, pH-responsive, sustained release properties. All of these characteristics would provide a robust multifunctional theranostic platform for visualized glioma therapy.

Pigs may need to be exploited as xenotransplantation donors due to the shortage of human organs, tissues and cells. Porcine endogenous retroviruses (PERVs) are a significant obstacle to xenotransplantation because they can infect human cells in vitro and have the potential for transmission of unexpected pathogens to humans. In this research. 101 pigs, including four commercial breeds (23 Berkshire, 13 Duroc, 22 Landrace and 14 Yorkshire pigs), one native breed (19 Korean native pigs) and one miniature breed (10 NIH miniature pigs) were used to investigate insertional variations for 11 PERV loci (three PERV-A, six PERV-B and two PERV-C). Over 60% of the pigs harbored one PERV-A (907F8) integration and five PERV-B (B3-3G, B3-7G, 742H1, 1155D9 and 465D1) integrations. However, two PERV-A loci (A1-6C and 1347C1) and one PERV-B locus (B3-7F) were absent in Duroc pigs. Moreover, two PERV-C loci (C2-6C and C4-2G) only existed in Korean native pigs and NIH miniature pigs. The results suggest that PERV insertional variations differ among pig breeds as well as among individuals within a breed. Also, the results presented here can be used for the selection of animals that do not have specific PERV integration for xenotransplantation research.

The cAMP receptor protein (CRP) requires cAMP for an allosteric change and regulates more than 150 genes in Escherichia coli. In this study, the modular half of cAMP receptor protein was used to investigate the allosteric signal transmission pathway induced by cAMP binding. The activation of CRP upon cAMP binding is indicated to be realignment of the two subunits within the CRP dimer. The interaction of loop 3 and Phe136 do not involve in signal transmission.

The cyclic AMP receptor protein (CRP) from Escherichia coli regulates the expression of a large number of genes. In this work, CRP has been overexpressed, purified and digested by subtilisin and chymotrypsin. The fragments S-CRP (digested by subtilisin) and CH-CRP (digested by chymotrypsin) have been purified and crystallized. Crystals of S-CRP diffracted to 2.0 A resolution and belonged to space group P2(1), with unit-cell parameters a = 59.7, b = 75.1, c = 128.3 A, beta = 91.5 degrees . Crystals of CH-CRP diffracted to 2.8 A resolution and belonged to space group P222, with unit-cell parameters a = 45.8, b = 60.9, c = 205.6 A.

Protein–nanoparticle interactions are important in biomedical applications of nanoparticles and for growing biosafety concerns about nanomaterials. In this study, the interactions of four plasma proteins, human serum albumin (HSA), myoglobin (MB), hemoglobin (HB), and trypsin (TRP), with Au and Ag nanoparticles were investigated by FT-IR spectroscopy. The secondary structure of thio-proteins changed with time during incubation with Au and Ag nanoparticles, but the secondary structures of non-thio-proteins remained unchanged. The incubation time for structural changes depended on the sulfur–metal bond energy; the stronger the sulfur–metal energy, the less the time needed. H/D exchange experiments revealed that protein–NP complexes with thio-proteins were less dynamic than free proteins. No measurable dynamic differences were found between free non-thio-proteins and the protein–Au (or Ag) nanoparticle complex. Therefore, the impact of covalent bonds on the protein structure is greater than that of the electrostatic force.

The tumor necrosis factor receptors (TNFRs) play essential roles in innate and adaptive immunity. Depending on conditions, TNFR induces multiple cell fates including cell survival, cell apoptosis, and cell programmed necrosis. Here, we review recent progress in structural studies of the TNFR signaling pathway. The structural basis for the high order signal complexes, including the DISC, ripoptosome, necrosome, and RIP3/MLKL complex, may provide novel insights for understanding the biophysical principles of cell signaling cascades.

The mosquito-borne West Nile (WNV) and dengue 2 (DEN2V) viruses and tick-borne Langat (LGTV) and Omsk hemorrhagic fever (OHFV) viruses are arthropod-borne flaviviruses (family Flaviviridae, genus Flavivirus). These viruses are quite similar at both the nucleotide and amino acid level, yet they are very divergent in their biological properties and in the diseases they cause. The objective of this study was to examine the putative receptor-binding domains of the flaviviruses, the envelope (E) protein domain III (D3), which assume very similar structures either as part of the whole envelope protein or as individual entities, and to define the biophysical properties that distinguish among these viruses. Circular dichroism and Fourier transform infrared spectroscopy were employed to monitor the solution structure of these proteins. While the spectroscopic results found that the D3 from each of these viruses is composed of either beta-sheets or beta-turns, which is consistent with X-ray crystal data for tick-borne encephalitis and dengue viruses, these results reveal that recombinant D3s (rED3s) derived from tick-borne flaviviruses (LGT-rED3 and OHF-rED3) were similar to each other, while those from mosquito-borne flaviviruses (WN-rED3 and DEN-rED3) were similar to each other yet distinct from rED3 of the tick-borne viruses. Protein dynamic studies probed by fluorescence quenching and hydrogen/deuterium exchange found that the rED3s are dynamic entities. The tick-borne proteins again exhibit very similar dynamic properties, which are different from the mosquito-borne proteins. The WN-rED3 is significantly less stable than the other three rED3s. Overall, these differences in biophysical properties correlate with biological properties of these viruses that tick-borne flaviviruses are more stable than mosquito-borne flaviviruses.

The Escherichia coli cyclic AMP receptor protein (CRP) is a prokaryotic global transcription activator protein that controls the expression of many different genes. Wild-type CRP can bind to special DNA sequences in the presence of cAMP. The substitution of Asp53 by His results in the CRP* phenotype, which does not require exogenous cAMP. In the present study, the D53H CRP mutant was overexpressed, purified and crystallized. cAMP-free D53H CRP crystals were obtained and diffracted to a resolution of 2.9 A. Based on the systematic absences of the crystals, the space group is likely to be P2(1)2(1)2(1), with unit-cell parameters a = 76.66, b = 152.14, c = 176.11 A. The asymmetric unit was confirmed to contain four protein dimers, with a Matthews coefficient of 2.71 A(3) Da(-1) and a solvent content of 54.68%.

The Escherichia coli cyclic AMP receptor protein (CRP) is a well known transcription activator protein. In this study, CRP was overexpressed, purified and cocrystallized with cAMP and a 38 bp full-length double-stranded DNA fragment. The full-length segment differed from the half-site fragments used in previous crystallization experiments and is more similar to the environment in vivo. CRP-cAMP-DNA crystals were obtained and diffracted to 2.9 Å resolution. The crystals belonged to space group P3121, with unit-cell parameters a = b = 76.03, c = 144.00 Å. The asymmetric unit was found to contain one protein molecule and half a 38 bp full-length double-stranded DNA fragment, with a Matthews coefficient of 2.62 Å(3) Da(-1) and a solvent content of 53.14%.

Multiplex gene assay is a valuable molecular tool not only in academic science but also in clinical diagnostics. Multiplex PCR assays, DNA microarrays, and various nanotechnology-based methods are examples of major techniques developed for analyzing multiple genes; none of these, however, are suitable for point-of-care diagnostics, especially in resource-limited settings. In this report, we describe an octopus-like multiplex microfluidic loop-mediated isothermal amplification (mμLAMP) assay for the rapid analysis of multiple genes in the point-of-care format and provide a robust approach for predicting viruses. This assay with the ability of analyzing multiple genes qualitatively and quantitatively is highly specific, operationally simple, and cost/time-effective with the detection limit of less than 10 copies/μL in 2 μL quantities of sample within 0.5 h. We successfully developed a mμLAMP chip for differentiating three human influenza A substrains and identifying eight important swine viruses.

The recombined ricin A chain protein (RTA) was transported into living cells by multiwalled carbon nanotubes (MWNTs) as a cellular carrier, as performed by natural ricin B chain protein (RTB). The conjugate of the toxin protein RTA and MWNT was found to translocate to the cytoplasm of various cell lines and performed biological functions, as evidenced by the induction of cell death. The delivery of RTA into the cells via nanotube carriers was directly visualized by transmission electron microscopy (TEM) and confocal microscopy. About three times higher cell death rates for L-929, HL7702, MCF-7, HeLa and COS-7 cells were demonstrated induced by MWNT-RTA conjugates, compared to those achieved by RTA alone. Especially for HeLa cells, the cell mortality reached approximately 75%. In addition, obvious selective destruction of cancer cells was achieved by coupling MWNTs-RTA-HER2, which selectively recognize the HER2/neu receptor on certain breast cancer cells. This is the first example of recombined protein toxin (RTA)-induced targeting destruction for tumor cells via carbon nanotube molecular transporters. The transporting capabilities of carbon nanotubes combined with functional proteins may open exciting new venues for drug delivery and cancer therapy.

Resolution of the 3D structures and IgE epitopes of allergens may identify common or conserved features of allergens. Jun a 1, the predominant allergen in mountain cedar pollen, was chosen as a model for identifying common structural and functional features among a group of plant allergens. In this study, synthetic, overlapping peptides of Jun a 1 and sera from patients allergic to mountain cedar pollen were used to identify linear epitopes. A 3D model of Jun a 1 was produced using the Bacillus subtiles pectate lyase (PL) as a template and validated with biophysical measurements. This allowed mappings of four IgE binding sites on Jun a 1. Two of the epitopes mapped to turns or loops on the surface of the model structure. The other two epitopes mapped to the β-sheet region, homologous to the catalytic site of PL. This region of Jun a 1 is highly conserved in the group 1 allergens from other cedar trees as well as microbial PLs. The finding that two out of three major IgE epitopes map to highly conserved catalytic regions of group 1 cedar allergens may help to explain the high degree of cross-reactivity between cedar pollen allergens and might represent a pattern of reactivity common to other allergens with catalytic activity.