The rise of baobab trees in Madagascar.

The baobab trees (genus Adansonia) have attracted tremendous attention because of their striking shape and distinctive relationships with fauna1. These spectacular trees have also influenced human culture, inspiring innumerable arts, folklore and traditions. Here we sequenced genomes of all eight extant baobab species and argue that Madagascar should be considered the centre of origin for the extant lineages, a key issue in their evolutionary history2,3. Integrated genomic and ecological analyses revealed the reticulate evolution of baobabs, which eventually led to the species diversity seen today. Past population dynamics of Malagasy baobabs may have been influenced by both interspecific competition and the geological history of the island, especially changes in local sea levels. We propose that further attention should be paid to the conservation status of Malagasy baobabs, especially of Adansonia suarezensis and Adansonia grandidieri, and that intensive monitoring of populations of Adansonia za is required, given its propensity for negatively impacting the critically endangered Adansonia perrieri.

Uncoupling transcription and translation through miRNA-dependent poly(A) length control in haploid male germ cells.

As one of the post-transcriptional regulatory mechanisms, uncoupling of transcription and translation plays an essential role in development and adulthood physiology. However, it remains elusive how thousands of mRNAs get translationally silenced while stability is maintained for hours or even days before translation. In addition to oocytes and neurons, developing spermatids display significant uncoupling of transcription and translation for delayed translation. Therefore, spermiogenesis represents an excellent in vivo model for investigating the mechanism underlying uncoupled transcription and translation. Through full-length poly(A) deep sequencing, we discovered dynamic changes in poly(A) length through deadenylation and re-polyadenylation. Deadenylation appeared to be mediated by microRNAs (miRNAs), and transcripts with shorter poly(A) tails tend to be sequestered into ribonucleoprotein (RNP) granules for translational repression and stabilization. In contrast, re-polyadenylation might allow for translocation of the translationally repressed transcripts from RNP granules to polysomes. Overall, our data suggest that miRNA-dependent poly(A) length control represents a previously unreported mechanism underlying uncoupled translation and transcription in haploid male mouse germ cells.

Improving the RNA velocity approach with single-cell RNA lifecycle (nascent, mature and degrading RNAs) sequencing technologies.

We presented an experimental method called FLOUR-seq, which combines BD Rhapsody and nanopore sequencing to detect the RNA lifecycle (including nascent, mature, and degrading RNAs) in cells. Additionally, we updated our HIT-scISOseq V2 to discover a more accurate RNA lifecycle using 10x Chromium and Pacbio sequencing. Most importantly, to explore how single-cell full-length RNA sequencing technologies could help improve the RNA velocity approach, we introduced a new algorithm called 'Region Velocity' to more accurately configure cellular RNA velocity. We applied this algorithm to study spermiogenesis and compared the performance of FLOUR-seq with Pacbio-based HIT-scISOseq V2. Our findings demonstrated that 'Region Velocity' is more suitable for analyzing single-cell full-length RNA data than traditional RNA velocity approaches. These novel methods could be useful for researchers looking to discover full-length RNAs in single cells and comprehensively monitor RNA lifecycle in cells.

Removal of polycyclic aromatic hydrocarbons (PAHs) and the response of indigenous bacteria in highly contaminated aged soil after persulfate oxidation.

Integrated chemical-biological treatment is a promising alternative to remove PAHs from contaminated soil, wherein indigenous bacteria is the key factor for the biodegradation of residual PAHs after the application of chemical oxidation. However, systematical study on the impact of persulfate (PS) oxidation on indigenous bacteria as well as PAHs removal is still scarce. In this study, the influences of different PS dosages (1%, 3%, 6%, and 10% [w/w]), as well as various activation methods (native iron, H2O2, alkaline, ferrous iron, and heat) on PAHs removal and indigenous bacteria in highly contaminated aged soil were investigated. Apparent degradation of PAHs in the soil treated with PS oxidation was observed, and the removal efficiency of total PAHs in the soil ranged from 38.28% to 79.97%. The removal efficiency of total PAHs in the soil increased with increasing consumption of PS. However, the bacterial abundance in soil was negatively affected following oxidation for all of the treatments added with PS, with bacterial abundance in the soil decreased by 0.89-2.93 orders of magnitude compared to the untreated soil. Moreover, the number of total bacteria in the soil decreased as PS consumption increased. Different PS activation methods and PS dosages exhibited different influences on the bacterial community composition. Bacteria capable of degrading PAHs under anoxic conditions were composed predominantly by Proteobacteria and Firmicutes. The total amount of Proteobacteria and Firmicutes also decreased with increasing consumption of PS. The results of this study provide important insight for the design of PAHs contaminated soil remediation projects.

iTRAQ-based comparative proteomic analysis of differentially expressed proteins in Rhodococcus sp. BAP-1 induced by fluoranthene.

To reveal the molecular mechanism at the level of regulation of proteins in Rhodococcus sp. BAP-1 induced by fluoranthene comparative proteomic analysis was performed on proteins extracted from fluoranthene-exposed cells on 1 d, 3 d, 6 d and 8 d compared with control cells using isobaric tags for relative and absolute quantization (iTRAQ) labeling and LC-MS/MS analysis to access differentially expressed proteins. As a result, we detected a total of 897 significantly differentially expressed proteins, including 30 shared proteins in four comparison clusters. We were able to short-list 190, 329, 101 and 90 proteins that were over-represented, and 394, 234, 65 and 49 under-represented proteins, in 1d/control, 3d/control, 6d/control and 8d/control comparisons, respectively. Functional analysis relied on Clusters of Orthologous Groups (COG), gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) revealed that fluoranthene significantly altered the expression of proteins involved in metabolic and biosynthesis processes. Furthermore, BAP-1 up-regulates aldehyde dehydrogenase, cytochrome c oxidase, and oligopeptide transport ATP-binding protein, while down-regulates several other proteins in order to adapt to fluoranthene exposure. These findings provide important clues to reveal fluoranthene degradation mechanism in BAP-1 and promote its bioremediation applications.

A lateral flow assay for the determination of human tetanus antibody in whole blood by using gold nanoparticle labeled tetanus antigen.

The authors describe a lateral flow assay (LFA) for the antibody against the infectious bacterium Clostridium tetani. Gold nanoparticles (AuNPs) were linked to tetanus antigen and are captured in the test line via the formation of a sandwich structure composed of AuNP-labeled tetanus antigen, tetanus antibody, and tetanus antigen. This leads to the formation of a characteristic red line due to the accumulation of AuNPs. The formation of the color line allows for a highly sensitive and selective detection of tetanus antibody, both with bare eyes and by smartphone-based quantitative analysis. This assay offers a wide detection range from 0 to 0.5 IU·mL-1 and has a linear relationship from 0.01 to 0.1 IU·mL-1 with an experimental detection limit of 0.01 IU·mL-1. This assay is simple, fast, inexpensive and highly selective. When applied to the detection of tetanus antibody in spiked whole blood, it provided reliable results that compared well to those obtained with a commercial ELISA kit. Graphical abstract ᅟ.

Label-free and rapid detection of ATP based on structure switching of aptamers.

In this work, an aptamer-based fluorescent strategy for label-free detection of ATP was developed by using Thioflavin T (ThT) as a fluorescence indicator, which can specifically bind with G-quadruplex DNAs to generate enhanced fluorescence intensity. In the absence of ATP, the folded structure of ATP aptamer allows the intercalation of ThT to produce strong fluorescence signal. However, upon ATP binding to the aptamer where ThT intercalated, the conformational change or distortion of the aptamer is large enough to cause much less intercalation of ThT and consequently drastic suppression of the fluorescence intensity. As such, the concentration of ATP could be identified very easily by observing fluorescence changes of this sensing system. This label-free assay could be accomplished very easily and quickly with a "mix-and-detect" detection method and exhibits high sensitivity to ATP with a detection limit of 33 nM in a wide range of 0.1-1000 µM. Furthermore, this proposed method is capable of detecting ATP in human serum and cell extracts. This method offers several advantages such as simplicity, rapidity, low cost, good stability and excellent selectivity, which make it hold great potential for the detection of ATP in bioanalytical and biological studies.

Neutrophil CD64 combined with PCT, CRP and WBC improves the sensitivity for the early diagnosis of neonatal sepsis.

BACKGROUND: The aim of this study was to determine whether neutrophil CD64 (nCD64) combined with procalcitonin (PCT), C-reactive protein (CRP) and white blood cell count (WBC) can increase the sensitivity and accuracy of neonatal sepsis diagnosis. METHODS: The serum levels of nCD64, CRP, PCT and WBC were detected in 60 patients with neonatal sepsis and 60 patients with non-sepsis. Sensitivity, specificity, positive and negative predictive values, receiver operating characteristic (ROC) area under the curve (AUC), and logistic regression analysis were performed to evaluate the diagnostic value of these markers on neonatal sepsis. RESULTS: Serum levels of nCD64, PCT, CRP and WBC were higher in the sepsis group than non-sepsis group (p<0.001). The sensitivities of nCD64, PCT, CRP and WBC at the recommended cut-off level for all infants were 79.5%, 68.2%, 38.6% and 52.3%, respectively. The best combination was nCD64 and PCT, which obtained sensitivity of 90.9%, largest AUC of 0.922, and a negative predictive value of 89.2%. However by using an optimal cut-off value, the sensitivities of all four biomarkers for the diagnosis of neonatal sepsis were increased to 95.5%. Except for WBC, the birth weight and gestational age had no effects on the diagnostic value of these serum biomarkers. CONCLUSIONS: nCD64 and PCT are better diagnostic biomarkers for early diagnosis of neonatal sepsis as compared to CRP. With the help of optimal cut-off value based on ROC curve and logistic regression analysis, the combination of these biomarkers could improve the sensitivity for the diagnosis of suspected late-onset neonatal sepsis based on common serum biomarkers.

A Turn on ESIPT Probe for Rapid and Ratiometric Fluorogenic Detection of Hg(2+) and its Application in Live-Cell Imaging.

A probe based on 2-(2'-hydroxyphenyl) benzothiazole (HBT) and thiophosphate has been synthesized and used for the ratiometric detection of Hg(2+). The probe was designed in such a way that the excited state intramolecular proton transfer (ESIPT) of the HBT moiety get blocked. The probe exhibited a strong fluorescence enhancement upon addition of Hg(2+) while showing almost no response to other cations in CH3CN/HEPES buffer solution. The probe exhibited fast selectivity towards Hg(2+) and could be completed in 1 min. Fluorescence imaging experiments of Hg(2+) in living TE-1 cells demonstrated its value of practical applications in biological systems.

Dihydro-Resveratrol Ameliorates Lung Injury in Rats with Cerulein-Induced Acute Pancreatitis.

Acute pancreatitis is an inflammatory process originated in the pancreas; however, it often leads to systemic complications that affect distant organs. Acute respiratory distress syndrome is indeed the predominant cause of death in patients with severe acute pancreatitis. In this study, we aimed to delineate the ameliorative effect of dihydro-resveratrol, a prominent analog of trans-resveratrol, against acute pancreatitis-associated lung injury and the underlying molecular actions. Acute pancreatitis was induced in rats with repetitive injections of cerulein (50 µg/kg/h) and a shot of lipopolysaccharide (7.5 mg/kg). By means of histological examination and biochemical assays, the severity of lung injury was assessed in the aspects of tissue damages, myeloperoxidase activity, and levels of pro-inflammatory cytokines. When treated with dihydro-resveratrol, pulmonary architectural distortion, hemorrhage, interstitial edema, and alveolar thickening were significantly reduced in rats with acute pancreatitis. In addition, the production of pro-inflammatory cytokines and the activity of myeloperoxidase in pulmonary tissues were notably repressed. Importantly, nuclear factor-kappaB (NF-κB) activation was attenuated. This study is the first to report the oral administration of dihydro-resveratrol ameliorated acute pancreatitis-associated lung injury via an inhibitory modulation of pro-inflammatory response, which was associated with a suppression of the NF-κB signaling pathway.

Study on Calculation Method of Bending Performance of Concrete Sandwich Composite Slab.

In order to explore the flexural behavior of a concrete sandwich panel under concentrated boundary conditions, based on Kirachhoff's elastic thin plate theory in this paper, the geometric deformation, physical conditions, and equilibrium relationship of a sandwich panel are deduced by constructing the layered analysis model of the sandwich panel, the basic differential equation of the flexural deformation of the concrete sandwich thin plate is obtained, and the mathematical expression of the internal force and displacement under the boundary condition of concentrated support is given. Combined with an engineering example, the proposed calculation method is verified. The results show that, in the arrangement of reliable connectors for concrete sandwich panels, the concrete wythes bear the load while the contribution of the core layer to the bending capacity of the structure can be ignored. When subjected to a laterally distributed load, the sandwich panel mainly experiences out-of-plane bending deformation, and the bending normal stress in the concrete panel layer shows a linear non-uniform distribution along the thickness direction of the panel. The bending deformation performance and bearing efficiency of a concrete sandwich slab with the change in concentrated support position have significant effects, and the load transfer efficiency can be improved by optimizing the arrangement of supports. Except for small local areas near the supports, the bending stress distribution and deformation behavior of the concrete sandwich panel can be accurately analyzed by the calculation method established in this paper.

Study on the accelerated biodegradation of PAHs in subsurface soil via coupled low-temperature thermally treatment and electron acceptor stimulation based on metagenomic sequencing.

In situ anoxic bioremediation is a sustainable technology to remediate PAHs contaminated soils. However, the limited degradation rate of PAHs under anoxic conditions has become the primary bottleneck hindering the application of this technology. In this study, coupled low-temperature thermally treatment (<50 °C) and EA biostimulation was used to enhance PAH removal. Anoxic biodegradation of PAHs in soil was explored in microcosms in the absence and presence of added EAs at 3 temperatures (15 °C, 30 °C, and 45 °C). The influence of temperature, EA, and their interaction on the removal of PAHs were identified. A PAH degradation model based on PLSR analysis identified the importance and the positive/negative role of parameters on PAH removal. Soil archaeal and bacterial communities showed similar succession patterns, the impact of temperature was greater than that of EA. Soil microbial community and function were more influenced by temperature than EAs. Close and frequent interactions were observed among soil bacteria, archaea, PAH-degrading genes and methanogenic genes. A total of 15 bacterial OTUs, 1 PAH-degrading gene and 2 methanogenic genes were identified as keystones in the network. Coupled low-temperature thermally treatment and EA stimulation resulted in higher PAH removal efficiencies than EA stimulation alone and low-temperature thermally treatment alone.

Mitophagy in Alzheimer's Disease: A Bibliometric Analysis from 2007 to 2022.

Background: The investigation of mitophagy in Alzheimer's disease (AD) remains relatively underexplored in bibliometric analysis. Objective: To delve into the progress of mitophagy, offering a comprehensive overview of research trends and frontiers for researchers. Methods: Basic bibliometric information, targets, and target-drug-clinical trial-disease extracted from publications identified in the Web of Science Core Collection from 2007 to 2022 were assessed using bibliometric software. Results: The study encompassed 5,146 publications, displaying a consistent 16-year upward trajectory. The United States emerged as the foremost contributor in publications, with the Journal of Alzheimer's Disease being the most prolific journal. P. Hemachandra Reddy, George Perry, and Xiongwei Zhu are the top 3 most prolific authors. PINK1 and Parkin exhibited an upward trend in the last 6 years. Keywords (e.g., insulin, aging, epilepsy, tauopathy, and mitochondrial quality control) have recently emerged as focal points of interest within the past 3 years. "Mitochondrial dysfunction" is among the top terms in disease clustering. The top 10 drugs/molecules (e.g., curcumin, insulin, and melatonin) were summarized, accompanied by their clinical trials and related targets. Conclusions: This study presents a comprehensive overview of the mitophagy research landscape in AD over the past 16 years, underscoring mitophagy as an emerging molecular mechanism and a crucial focal point for potential drug in AD. This study pioneers the inclusion of targets and their correlations with drugs, clinical trials, and diseases in bibliometric analysis, providing valuable insights and inspiration for scholars and readers of JADR interested in understanding the potential mechanisms and clinical trials in AD.

Comparative proteomics reveals that fatty acid metabolism is involved in myocardial adaptation to chronic hypoxic injury.

Congenital heart disease (CHD) is the most serious form of heart disease, and chronic hypoxia is the basic physiological process underlying CHD. Some patients with CHD do not undergo surgery, and thus, they remain susceptible to chronic hypoxia, suggesting that some protective mechanism might exist in CHD patients. However, the mechanism underlying myocardial adaptation to chronic hypoxia remains unclear. Proteomics was used to identify the differentially expressed proteins in cardiomyocytes cultured under hypoxia for different durations. Western blotting assays were used to verify protein expression. A Real-Time Cell Analyzer (RTCA) was used to analyze cell growth. In this study, 3881 proteins were identified by proteomics. Subsequent bioinformatics analysis revealed that proteins were enriched in regulating oxidoreductase activity. Functional similarity cluster analyses showed that chronic hypoxia resulted in proteins enrichment in the mitochondrial metabolic pathway. Further KEGG analyses found that the proteins involved in fatty acid metabolism, the TCA cycle and oxidative phosphorylation were markedly upregulated. Moreover, knockdown of CPT1A or ECI1, which is critical for fatty acid degradation, suppressed the growth of cardiomyocytes under chronic hypoxia. The results of our study revealed that chronic hypoxia activates fatty acid metabolism to maintain the growth of cardiomyocytes.

Identification of Blood Biomarkers Related to Energy Metabolism and Construction of Diagnostic Prediction Model Based on Three Independent Alzheimer's Disease Cohorts.

Background: Blood biomarkers are crucial for the diagnosis and therapy of Alzheimer's disease (AD). Energy metabolism disturbances are closely related to AD. However, research on blood biomarkers related to energy metabolism is still insufficient. Objective: This study aims to explore the diagnostic and therapeutic significance of energy metabolism-related genes in AD. Methods: AD cohorts were obtained from GEO database and single center. Machine learning algorithms were used to identify key genes. GSEA was used for functional analysis. Six algorithms were utilized to establish and evaluate diagnostic models. Key gene-related drugs were screened through network pharmacology. Results: We identified 4 energy metabolism genes, NDUFA1, MECOM, RPL26, and RPS27. These genes have been confirmed to be closely related to multiple energy metabolic pathways and different types of T cell immune infiltration. Additionally, the transcription factors INSM2 and 4 lncRNAs were involved in regulating 4 genes. Further analysis showed that all biomarkers were downregulated in the AD cohorts and not affected by aging and gender. More importantly, we constructed a diagnostic prediction model of 4 biomarkers, which has been validated by various algorithms for its diagnostic performance. Furthermore, we found that valproic acid mainly interacted with these biomarkers through hydrogen bonding, salt bonding, and hydrophobic interaction. Conclusions: We constructed a predictive model based on 4 energy metabolism genes, which may be helpful for the diagnosis of AD. The 4 validated genes could serve as promising blood biomarkers for AD. Their interaction with valproic acid may play a crucial role in the therapy of AD.

Lateralized brunt of sleep deprivation on white matter injury in a rat model of Alzheimer's disease.

Sleep disturbance is a recognized risk factor for Alzheimer's disease (AD), but the underlying micro-pathological evidence remains limited. To bridge this gap, we established an amyloid-ß oligomers (AßO)-induced rat model of AD and subjected it to intermittent sleep deprivation (SD). Diffusion tensor imaging (DTI) and transmission electron microscopy were employed to assess white matter (WM) integrity and ultrastructural changes in myelin sheaths. Our findings demonstrated that SD exacerbated AßO-induced cognitive decline. Furthermore, we found SD aggravated AßO-induced asymmetrical impairments in WM, presenting with reductions in tract integrity observed in commissural fibers and association fasciculi, particularly the right anterior commissure, right corpus callosum, and left cingulum. Ultrastructural changes in myelin sheaths within the hippocampus and corpus callosum further confirmed a lateralized effect. Moreover, SD worsened AßO-induced lateralized disruption of the brain structural network, with impairments in critical nodes of the left hemisphere strongly correlated with cognitive dysfunction. This work represents the first identification of a lateralized impact of SD on the mesoscopic network and cognitive deficits in an AD rat model. These findings could deepen our understanding of the complex interplay between sleep disturbance and AD pathology, providing valuable insights into the early progression of the disease, as well as the development of neuroimaging biomarkers for screening early AD patients with self-reported sleep disturbances. Enhanced understanding of these mechanisms may pave the way for targeted interventions to alleviate cognitive decline and improve the quality of life for individuals at risk of or affected by AD.

Fusarium pseudograminearum biomass and toxin accumulation in wheat tissues with and without Fusarium crown rot symptoms.

Fusarium crown rot (FCR) is an important and devastating disease of wheat (Triticum aestivum) caused by the fungus Fusarium pseudograminearum and related pathogens. Using two distinct susceptible cultivars, we investigated the isolation frequencies of F. pseudograminearum and quantified its biomass accumulation and the levels of the associated toxins deoxynivalenol (DON) and DON-3-glucoside (D3G) in inoculated field-grown wheat plants. We detected F. pseudograminearum in stem, peduncle, rachis, and husk tissues, but not in grains, whereas DON and D3G accumulated in stem, rachis, husk, and grain tissues. Disease severity was positively correlated with the frequency of pathogen isolation, F. pseudograminearum biomass, and mycotoxin levels. The amount of F. pseudograminearum biomass and mycotoxin contents in asymptomatic tissue of diseased plants were associated with the distance of the tissue from the diseased internode and the disease severity of the plant. Thus, apparently healthy tissue may harbor F. pseudograminearum and contain associated mycotoxins. This research helps clarify the relationship between F. pseudograminearum occurrence, F. pseudograminearum biomass, and mycotoxin accumulation in tissues of susceptible wheat cultivars with or without disease symptoms, providing information that can lead to more effective control measures.

Spatial chromatin accessibility sequencing resolves high-order spatial interactions of epigenomic markers.

As the genome is organized into a three-dimensional structure in intracellular space, epigenomic information also has a complex spatial arrangement. However, most epigenetic studies describe locations of methylation marks, chromatin accessibility regions, and histone modifications in the horizontal dimension. Proper spatial epigenomic information has rarely been obtained. In this study, we designed spatial chromatin accessibility sequencing (SCA-seq) to resolve the genome conformation by capturing the epigenetic information in single-molecular resolution while simultaneously resolving the genome conformation. Using SCA-seq, we are able to examine the spatial interaction of chromatin accessibility (e.g. enhancer-promoter contacts), CpG island methylation, and spatial insulating functions of the CCCTC-binding factor. We demonstrate that SCA-seq paves the way to explore the mechanism of epigenetic interactions and extends our knowledge in 3D packaging of DNA in the nucleus.

Whole-genome Sequencing Reveals Autooctoploidy in Chinese Sturgeon and Its Evolutionary Trajectories.

The order Acipenseriformes, which includes sturgeons and paddlefishes, represents "living fossils" with complex genomes that are good models for understanding whole-genome duplication (WGD) and ploidy evolution in fishes. Here, we sequenced and assembled the first high-quality chromosome-level genome for the complex octoploid Acipenser sinensis (Chinese sturgeon), a critically endangered species that also represents a poorly understood ploidy group in Acipenseriformes. Our results show that A. sinensis is a complex autooctoploid species containing four kinds of octovalents (8n), a hexavalent (6n), two tetravalents (4n), and a divalent (2n). An analysis taking into account delayed rediploidization reveals that the octoploid genome composition of Chinese sturgeon results from two rounds of homologous WGDs, and further provides insights into the timing of its ploidy evolution. This study provides the first octoploid genome resource of Acipenseriformes for understanding ploidy compositions and evolutionary trajectories of polyploid fishes.

Fluorescence protection assay: a novel homogeneous assay platform toward development of aptamer sensors for protein detection.

Development of novel aptamer sensor strategies for rapid and selective assays of protein biomarkers plays crucial roles in proteomics and clinical diagnostics. Herein, we have developed a novel aptamer sensor strategy for homogeneous detection of protein targets based on fluorescence protection assay. This strategy is based on our reasoning that interaction of aptamer with its protein target may dramatically increase steric hindrance, which protects the fluorophore, fluorescein isothiocyannate (FITC), labeled at the binding pocket from accessing and quenching by the FITC antibody. The aptamer sensor strategy is demonstrated using a model protein target of immunoglobulin E (IgE), a known biomarker associated with atopic allergic diseases. The results reveal that the aptamer sensor shows substantial (>6-fold) fluorescence enhancement in response to the protein target, thereby verifying the mechanism of fluorescence protection. Moreover, the aptamer sensor displays improved specificity to other co-existing proteins and a desirable dynamic range within the IgE concentration from 0.1 to 50 nM with a readily achieved detection limit of 0.1 nM. Because of great robustness, easy operation and scalability for parallel assays, the developed homogeneous fluorescence protection assay strategy might create a new methodology for developing aptamer sensors in sensitive, selective detection of proteins.