Design and calibration of the 6-DOF motion tracking system integrated on the Stewart parallel manipulator.

Accurate pose measurement is crucial for parallel manipulators (PM). This study designs a novel integrated 6-DOF motion tracking system to achieve precise online pose measurement. However, the presence of geometric errors introduces imperfections in the accuracy of the measured pose. Based on the displacement information of six grating rulers, measurement pose is obtained through forward kinematics. By comparing the measurement results with the actual pose information captured by stereo vision, measurement errors can be obtained. A closed-loop vector-based kinematic model and an error model are established, and then the geometric errors are identified with the least-squares method. Finally, the geometric calibration experiments are conducted, and the results show that the measurement accuracy has significantly improved, with the average position error decreasing from 3.148 mm to 0.036 mm, and the average orientation error is decreased from 0.225° to 0.022°.

Monocular vision-based dynamic calibration method for determining the sensitivities of low-frequency tri-axial vibration sensors.

Low-frequency vibrations exist widely in the natural environment and in human activities. Low-frequency tri-axial vibration sensors are enormously applied in the fields of seismic monitoring, building structure health monitoring, aerospace navigating, etc. Their sensitivity calibration accuracy directly determines whether their applications can work reliably. Although the laser interferometry recommended by the International Standardization Organization (ISO) is commonly used to achieve the vibration calibration, it suffers from the shortages of low-frequency range, high cost, low efficiency, and limited applicable environment. In this study, a novel monocular vision-based dynamic calibration method is proposed, which determines the whole sensitivities of tri-axial sensors by the monocular vision method to accurately measure the spatial input excitation. This method improves the calibration performance by eliminating the installation error and enhancing calibration efficiency via decreasing reinstallations. The experimental results compared with the laser interferometry demonstrate that the investigated method can obtain similar calibration accuracy in the range of 0.16-2 Hz with more efficiency. The corresponding maximum relative deviations of X-, Y-, and Z-axial sensitivities were approximately 2.5%, 1.8%, and 0.4%. In addition, the maximum relative standard deviation of the investigated method was only about 0.3% in this range.

RER-YOLO: improved method for surface defect detection of aluminum ingot alloy based on YOLOv5.

Aluminum ingot alloy is one of the commonly used materials in industrial production and intelligent manufacturing, whose quality directly affects the performance of aluminum processed products. Therefore, the inspection of surface defects of aluminum ingot alloy is extremely valuable for actual industrial engineering. Aiming at the issues of low detecting precision and the slowly processing rate thatexisted in the traditional target detection methods for aluminum ingot alloy dataset, the YOLOv5-based improvement model RER-YOLO is proposed. Firstly, the aluminum ingot alloy dataset is coped with the image pretreatment methods of rotation, translation, contrast and brightness transformations in a random combination so as to boost the capacity of generalization for model training. Secondly, a multi-scale characteristic extraction network block (Res2Net) is utilized to take the place of the C3 block in the previous YOLOv5 to augment the model's ability that can accurately extract rich features. Finally, an over-parameterization-based re-parameterized convolutional block is utilized in place of the 3×3 convolutional blocks in the Res2Net residual block and baseline model, enlarging the search space of the network and boosting the model's fitting ability while maintaining inference rate. The comparison experimental results demonstrate that the RER-YOLO reaches a mean average precision of 75.1% on the aluminum ingot alloy dataset, which is higher 4.9% than the conventional YOLOv5 and does not increase the inference delay. It also improves the detection accuracy by 12.7% for burr defects, which are fewer in number in the dataset and the defect features are difficult to extract. It can be seen that the presented model in this study has an important reference value towards detecting surface defects in aluminum ingot alloy.

Standard sine motion video-based virtual traceability method for monocular vision low-frequency vibration measurements.

The low-frequency vibration exists in building structures, mechanical devices, instrument manufacturing, and other fields, and is the key to modal analysis, steady-state control, and precision machining. At present, the monocular vision (MV) method has gradually become the primary choice to measure the low-frequency vibration because of its distinctive advantages in efficiency, non-contact, simplicity, flexibility, low cost, etc. Although many literature reports have demonstrated that this method has the capability to reach high measurement repeatability and resolution, its metrological traceability and uncertainty evaluation are difficult to be unified. In this study, a novel, to the best of our knowledge, virtual traceability method is presented to evaluate the measurement performance of the MV method for the low-frequency vibration. This presented method achieves traceability by adopting the standard sine motion videos and the precise position error correction model. Simulations and experiments confirm that the presented method can evaluate the amplitude and phase measurement accuracy of the MV-based low-frequency vibration in the frequency range from 0.01 to 20 Hz.

Stereo vision-based Kinematic calibration method for the Stewart platforms.

Accuracy is the most important index for the industrial applications of the Stewart platform, which can be guaranteed by the kinematic calibration method to improve the motion orbit performance of this platform. In order to improve the effectiveness of the least squares algorithm and the identified accuracy of the platform's geometric parameter errors, an applicab-le dimensionless error model based on the structural characteristics of the Stewart platform is investigated. Moreover, a novel stereo vision-based measurement method is proposed, which can measure the 6-degree-of-freedom (DOF) pose of the moving platform. On this basis, an identification simulation is schemed to validate the efficiency of the dimensionless error model, and the kinematic calibration experiment is carried out on a prototype. The experimental results demonstrate that the position error is decreased to 0.261 mm with an improved accuracy of 89.720%, the orientation error is decreased to 0.051° with an improved accuracy of 90.351%.

GdX/UBL4A specifically stabilizes the TC45/STAT3 association and promotes dephosphorylation of STAT3 to repress tumorigenesis.

Impaired phosphatase activity contributes to the persistent activation of STAT3 in tumors. Given that STAT family members with various or even opposite functions are often phosphorylated or dephosphorylated by the same enzymes, the mechanism for STAT3-specific dephosphorylation in cells remains largely unknown. Here, we report that GdX (UBL4A) promotes STAT3 dephosphorylation via mediating the interaction between TC45 (the nuclear isoform of TC-PTP) and STAT3 specifically. GdX stabilizes the TC45-STAT3 complex to bestow upon STAT3 an efficient dephosphorylation by TC45. Inasmuch, GdX suppresses tumorigenesis and tumor development by reducing the level of phospho-STAT3 (p-STAT3), whereas deletion of GdX results in a high level of p-STAT3 and accelerated colorectal tumorigenesis induced by AOM/DSS. Thus, GdX converts TC45, a nonspecific phosphatase, into a STAT3-specific phosphatase by bridging an association between TC45 and STAT3.

Visual encoder-based angle measurement method in low-frequency angular vibration calibration.

Angular vibration calibration is required to determine the sensitivity of sensors such as dynamic inclinometers, gyroscopes, and angular accelerometers, which are used for angular motion measurement in engineering applications. Additionally, the calibration performance depends on the accuracy of the angle measurement by laser interferometry or a circular grating (CG) method that is commonly used in vibration calibration. However, these methods usually own a complex and high-cost system or limited frequency and amplitude ranges. In this study, a novel, to the best of our knowledge, angle measurement method that combines a special visual encoder and an accurate angular position detection method is investigated; the method requires only a simple and flexible telecentric vision measurement system. Comparison experiments with the CG method demonstrate that the investigated method has the maximum measurement deviation of 0.0014° and 0.0138° for static angle measurement in the small-angle range and continuous full circle, respectively. The relative deviation of the angular vibration measurement in the range of 0.1-8 Hz with amplitudes 0-100° is less than 0.173%. Additionally, the relative deviation of calibrated sensitivity of a gyroscope by the investigated and CG methods is less than 0.096%.

High-Acceleration Vibration Calibration System Based on Phase-Locked Resonance Control.

In order to ensure the measurement accuracy of high-acceleration vibration sensors used in engineering applications, it is necessary to calibrate their key performance parameters at high acceleration. The high-acceleration vibration calibration system produces high-acceleration vibration by utilizing the resonance amplification principle; however, the resonance frequency of the resonant beam changes with increasing amplitude, affected by the influences of nonlinear and other factors. In this study, a phase-locked resonance tracking control method based on the phase resonance principle is proposed to accurately and quickly track the resonance frequency of the resonant beam, which can improve the accuracy and stability of resonance control. The resonant beam is able to produce stable vibration with an amplitude exceeding 7500 m/s2 by phase-locking and tracking the resonant frequency. A calibration system built with this method can provide stable vibration with an amplitude of 500-10,000 m/s2 in the range of 80-4000 Hz. Comparison experiments with the commonly used amplitude iteration amplification method demonstrate that the proposed method can give an acceleration stability control index of less than 0.5% and a resonance tracking time of less than 0.1 s.

Monocular vision-based low-frequency vibration calibration method with correction of the guideway bending in a long-stroke shaker.

Calibration is required to maximize the sensitivity and measurement accuracy of vibration sensors. In this study, a low-frequency vibration calibration method is proposed that is based on the concept of monocular vision. In this method, we employ a high-accuracy edge extraction method to extract the edges of sequential images so as to obtain the high calibration accuracy. However, the proposed method must rely on a long-stroke shaker to provide vibration excitation to the sensor, and the bending in the guideway caused by the mechanical processing reduces the calibration accuracy, especially at very low frequencies. The proposed setting compensates for the bending using an additional monocular vision technique to significantly improve the calibration accuracy. To validate the calibration accuracy of the proposed method, a comparison was conducted between results obtained via the laser interferometry, the Earth's gravitation method, and the proposed method when applied to calibrate the sensitivity of a tri-axial acceleration sensor at frequencies between 0.04 and 8 Hz. The results of the comparison showed the proposed method calibrated the sensor sensitivity with high accuracy and was able to accurately account for the bending when the frequency was lower than 0.3 Hz. In contrast, the calibration accuracy of the laser interferometry decreased because of the bending.

Testing of a MEMS Dynamic Inclinometer Using the Stewart Platform.

The micro-electro-mechanical system (MEMS) dynamic inclinometer integrates a tri-axis gyroscope and a tri-axis accelerometer for real-time tilt measurement. The Stewart platform has the ability to generate six degrees of freedom of spatial orbits. The method of applying spatial orbits to the testing of MEMS inclinometers is investigated. Inverse and forward kinematics are analyzed for controlling and measuring the position and orientation of the Stewart platform. The Stewart platform is controlled to generate a conical motion, based on which the sensitivities of the gyroscope, accelerometer, and tilt sensing are determined. Spatial positional orbits are also generated in order to obtain the tilt angles caused by the cross-coupling influence. The experiment is conducted to show that the tested amplitude frequency deviations of the gyroscope and tilt sensing sensitivities between the Stewart platform and the traditional rotator are less than 0.2 dB and 0.1 dB, respectively.

Bandpass-sampling-based heterodyne interferometer signal acquisition for vibration measurements in primary vibration calibration.

Heterodyne interferometers have been widely used for primary vibration calibration in recent years. Primary vibration calibration performance depends on the precision and real-time nature of vibration measurements, factors determined by the acquisition and demodulation of the heterodyne interferometer signal. This signal is commonly collected using the Nyquist sampling method, requiring devices with high sampling rates and large memories, or a sampling method using a mixer and low-pass filter-analog devices which may create time delays. This study proposes a novel bandpass sampling method that reduces sampling rate and storage capacity without generating time delays. To improve vibration measurement precision, an optimal sampling rate is designed to collect the heterodyne interferometer signal, and the collected signal is demodulated using the phase-unwrapping sine approximation method. The proposed method is compared with existing methods through simulated and experimental data. Experimental results show that the proposed method avoids time delays and high sampling rates, while providing high-precision vibration measurements.

A broad neutralizing nanobody against SARS-CoV-2 engineered from an approved drug.

SARS-CoV-2 infection is initiated by Spike glycoprotein binding to the human angiotensin-converting enzyme 2 (ACE2) receptor via its receptor binding domain. Blocking this interaction has been proven to be an effective approach to inhibit virus infection. Here we report the discovery of a neutralizing nanobody named VHH60, which was directly produced from an engineering nanobody library based on a commercialized nanobody within a very short period. VHH60 competes with human ACE2 to bind the receptor binding domain of the Spike protein at S351, S470-471and S493-494 as determined by structural analysis, with an affinity of 2.56 nM. It inhibits infections of both ancestral SARS-CoV-2 strain and pseudotyped viruses harboring SARS-CoV-2 wildtype, key mutations or variants at the nanomolar level. Furthermore, VHH60 suppressed SARS-CoV-2 infection and propagation 50-fold better and protected mice from death for twice as long as the control group after SARS-CoV-2 nasal infections in vivo. Therefore, VHH60 is not only a powerful nanobody with a promising profile for disease control but also provides evidence for a highly effective and rapid approach to generating therapeutic nanobodies.

Secretory expression and efficient purification of recombinant anthrax toxin lethal factor with full biological activity in E. coli.

Lethal factor (LF), a virulence factor of Bacillus anthracis, plays key roles in anthrax pathogenesis and host-pathogen interactions. The detailed mechanisms by which LF contributes to infection are still under investigation. While these studies require pure, homogeneous and reliable LF preparations, most methods reported for production of recombinant LF (rLF) in B. anthracis or Escherichia coli either are complicated or add extra residues to the protein. In this work, we modified our previous method by codon optimization and chromatograph workflow refinement and developed an improved strategy for efficient production of rLF from the periplasm of E. coli. We were able to obtain fully functional rLF with a purity above 95% and with a considerable yield of 5 mg/L. The preparation was characterized by SDS-PAGE, Western blot, and N-terminal sequencing, and the activity was validated by intoxication of macrophages and Fischer 344 rats. Our final product is suitable for most research involving drug development and mechanism analysis of anthrax pathogenesis.

Crystallization and preliminary X-ray analysis of the vWA domain of human anthrax toxin receptor 1.

The Gram-positive spore-forming bacterium Bacillus anthracis causes anthrax by secreting anthrax toxin, which consists of protective antigen (PA), lethal factor and oedema factor. Binding of PA to receptors triggers the multi-step process of anthrax toxin entry into target cells. Two distinct cellular receptors, ANTXR1 (also known as tumour endothelial marker 8; TEM8) and ANTXR2 (also known as capillary morphogenesis protein 2; CMG2), for anthrax toxin have been identified. Although the crystal structure of the extracellular von Willebrand factor A (vWA) domain of CMG2 has been reported, the difference between the vWA domains of TEM8 and CMG2 remains unclear because there are no structural data for the TEM8 vWA domain. In this report, the TEM8 vWA domain was expressed, purified and crystallized. X-ray diffraction data were collected to 1.8 Šresolution from a single crystal, which belonged to space group P1 with unit-cell parameters a=65.9, b=66.1, c=74.4 Å, α=63.7, ß=88.2, γ=59.9°.

The inhibition of the interaction between the anthrax toxin and its cellular receptor by an anti-receptor monoclonal antibody.

The high affinity binding of the anthrax protective antigen (PA) to one of its receptors, capillary morphogenesis protein 2 (CMG2), is essential for the intoxication process of anthrax toxin. To acquire novel research tools to study the PA-CMG2 interaction, we generated several anti-CMG2 monoclonal antibodies (MAbs). We demonstrated that one of the MAbs, 4B5, could inhibit PA-CMG2 binding and could also protect the sensitive cells against an anthrax lethal toxin challenge. We identified the epitope recognized by 4B5 and confirmed that the key residues of the epitope were the residues (119)YI-LK(125) of CMG2. Based on our results, we propose that 4B5 binds to the E122 pocket of CMG2 and interrupts the interaction between the pocket and the PA 2beta3-2beta4 loop. To our knowledge, this is the first report to illustrate that an anti-CMG2 antibody could inhibit the PA-CMG2 interaction and therefore interfere with the intoxication of anthrax toxin.

Deletion modification enhances anthrax specific immunity and protective efficacy of a hepatitis B core particle-based anthrax epitope vaccine.

Protective antigen (PA) is one of the major virulence factors of anthrax and is also the major constituent of the current anthrax vaccine. Previously, we found that the 2ß2-2ß3 loop of PA contains a dominant neutralizing epitope, the SFFD. We successfully inserted the 2ß2-2ß3 loop of PA into the major immunodominant region (MIR) of hepatitis B virus core (HBc) protein. The resulting fusion protein, termed HBc-N144-PA-loop2 (HBcL2), can effectively produce anthrax specific protective antibodies in an animal model. However, the protective immunity caused by HBcL2 could still be improved. In this research, we removed amino acids 79-81 from the HBc MIR of the HBcL2. This region was previously reported to be the major B cell epitope of HBc, and in keeping with this finding, we observed that the short deletion in the MIR not only diminished the intrinsic immunogenicity of HBc but also stimulated a higher titer of anthrax specific immunity. Most importantly, this deletion led to the full protection of the immunized mice against a lethal dose anthrax toxin challenge. We supposed that the conformational changes which occurred after the short deletion and foreign insertion in the MIR of HBc were the most likely reasons for the improvement in the immunogenicity of the HBc-based anthrax epitope vaccine.

Tumor endothelium marker-8 based decoys exhibit superiority over capillary morphogenesis protein-2 based decoys as anthrax toxin inhibitors.

Anthrax toxin is the major virulence factor produced by Bacillus anthracis. The toxin consists of three protein subunits: protective antigen (PA), lethal factor, and edema factor. Inhibition of PA binding to its receptors, tumor endothelium marker-8 (TEM8) and capillary morphogenesis protein-2 (CMG2) can effectively block anthrax intoxication, which is particularly valuable when the toxin has already been overproduced at the late stage of anthrax infection, thus rendering antibiotics ineffectual. Receptor-like agonists, such as the mammalian cell-expressed von Willebrand factor type A (vWA) domain of CMG2 (sCMG2), have demonstrated potency against the anthrax toxin. However, the soluble vWA domain of TEM8 (sTEM8) was ruled out as an anthrax toxin inhibitor candidate due to its inferior affinity to PA. In the present study, we report that L56A, a PA-binding-affinity-elevated mutant of sTEM8, could inhibit anthrax intoxication as effectively as sCMG2 in Fisher 344 rats. Additionally, pharmacokinetics showed that L56A and sTEM8 exhibit advantages over sCMG2 with better lung-targeting and longer plasma retention time, which may contribute to their enhanced protective ability in vivo. Our results suggest that receptor decoys based on TEM8 are promising anthrax toxin inhibitors and, together with the pharmacokinetic studies in this report, may contribute to the development of novel anthrax drugs.

[Establishment of RAW264.7 cell line stably expressing Mycobacterium tuberculosis protein ESAT-6].

For studying the effects of Mycobacterium tuberculosis secretory protein ESAT-6 on the related functions of macrophages, RAW264.7 cells were transfected with pEGFP-C1-ESAT-6 and pEGFP-C1 by liposome respectively. After screening with a high level of G418, the macrophage cell lines that stably expressed EGFP-ESAT-6 fusion protein or EGFP were established. The gene and protein expression levels were further analyzed by RT-PCR, fluorescence microscopy and Western blotting. The results indicated that the EGFP-ESAT6 fusion gene was integrated into the chromosome and the protein could be stably expressed in the selected macrophage cell line. These results gave us a tool for the future study in the mechanisms of ESAT-6 protein in modulating the macrophage cells.

The structure of tumor endothelial marker 8 (TEM8) extracellular domain and implications for its receptor function for recognizing anthrax toxin.

Anthrax toxin, which is released from the gram-positive bacterium Bacillus anthracis, is composed of three proteins: protective antigen (PA), lethal factor (LF), and edema factor (EF). PA binds a receptor on the surface of the target cell and further assembles into a homo-heptameric pore through which EF and LF translocate into the cytosol. Two distinct cellular receptors for anthrax toxin, TEM8/ANTXR1 and CMG2/ANTXR2, have been identified, and it is known that their extracellular domains bind PA with low and high affinities, respectively. Here, we report the crystal structure of the TEM8 extracellular vWA domain at 1.7 A resolution. The overall structure has a typical integrin fold and is similar to that of the previously published CMG2 structure. In addition, using structure-based mutagenesis, we demonstrate that the putative interface region of TEM8 with PA (consisting of residues 56, 57, and 154-160) is responsible for the PA-binding affinity differences between the two receptors. In particular, Leu56 was shown to be a key factor for the lower affinity of TEM8 towards PA compared with CMG2. Because of its high affinity for PA and low expression in normal tissues, an isolated extracellular vWA domain of the L56A TEM8 variant may serve as a potent antitoxin and a potential therapeutic treatment for anthrax infection. Moreover, as TEM8 is often over-expressed in tumor cells, our TEM8 crystal structure may provide new insights into how to design PA mutants that preferentially target tumor cells.

Fusion protein of Delta 27LFn and EFn has the potential as a novel anthrax toxin inhibitor.

PA-binding domain of LF (LFn) or PA-binding domain of EF (EFn) is the anthrax protective antigen (PA) binding domain of anthrax lethal factor (LF) or edema factor (EF). Here we show the development of a novel anthrax toxin inhibitor, fusion protein of N-terminal 27 amino acids deletion of LFn (Delta27LFn) and EFn. In a cell model of intoxication, fusion protein of Delta27LFn and EFn (Delta27LFn-EFn) was a 62-fold more potent toxin inhibitor than LFn or EFn, and this increased activity corresponded to a 39-fold higher PA-binding affinity by Biacore analysis. More importantly, Delta27LFn-EFn could protect the highly susceptible Fischer 344 rats from anthrax lethal toxin challenge. This work suggested that Delta27LFn-EFn has the potential as a candidate therapeutic agent against anthrax.