Nonlinear causal network learning via Granger causality based on extreme support vector regression.

For complex networked systems, based on the consideration of nonlinearity and causality, a novel general method of nonlinear causal network learning, termed extreme support vector regression Granger causality (ESVRGC), is proposed. The nonuniform time-delayed influence of the driving nodes on the target node is particularly considered. Then, the restricted model and the unrestricted model of Granger causality are, respectively, formulated based on extreme support vector regression, which uses the selected time-delayed components of system variables as the inputs of kernel functions. The nonlinear conditional Granger causality index is finally calculated to confirm the strength of a causal interaction. Generally, based on the simulation of a nonlinear vector autoregressive model and nonlinear discrete time-delayed dynamic systems, ESVRGC demonstrates better performance than other popular methods. Also, the validity and robustness of ESVRGC are also verified by the different cases of network types, sample sizes, noise intensities, and coupling strengths. Finally, the superiority of ESVRGC is successful verified by the experimental study on real benchmark datasets.

How Work Stress Impacts Emotional Outcomes of Chinese College Teachers: The Moderated Mediating Effect of Stress Mindset and Resilience.

Based on the job demands-resources model and conservation of resource theory, this study investigated 456 Chinese college teachers' work stress, stress mindset, resilience, emotional exhaustion, positive affect, and negative affect. The results of mediation analysis showed that resilience played a partial mediation role between work stress and emotional outcomes (emotional exhaustion, positive affect, and negative affect). Moreover, the results of a moderated mediation analysis showed that stress mindset moderated the relationship between work stress and resilience, and moderated the mediating effect of resilience between work stress and emotional outcomes (emotional exhaustion, positive affect, and negative affect). Specifically, work stress had a significant negative predictive effect on resilience when stress mindset is low (ß = -0.54, p < 0.001); work stress could also negatively predict resilience when the stress mindset is high (ß = -0.47, p < 0.001), but its effect decreased, and stress mindset negatively moderated the path between work stress and resilience. Finally, we discussed theoretical implications, practical implications, limitations, and future directions.

Analysis on the Efficacy of Bronchial Artery Chemoembolization Combined with 125I Seed Implantation in the Therapy of Advanced Non-Small-Cell Lung Cancer Based on the Medical Database.

Objective: To investigate the use and the efficacy of bronchial artery chemoembolization combined with 125I seed implantation in advanced non-small-cell lung cancer (NSCLC) therapy based on the medical database. Methods: A total of 102 patients with advanced NSCLC were randomly divided into two groups. The control group was treated with 125I seed implantation, and the observation group was treated with bronchial artery chemoembolization (BACE) combined with 125I seed implantation based on medical database. The clinical efficacy, carcinoembryonic antigen (CEA), cytokeratin 19 fragment antigen 21-1 (CYFRA21-1), glycan antigen 125 (CA125), peripheral blood CD3+, CD8+, CD4+/CD8+ T cells, insulin-like growth factor type 1 receptor (IGF-1R), S100 calcium-binding protein A2 (S100A2), long-term efficacy (time to disease progression, six-month survival rate, and one-year survival rate), and safety were then analyzed. Result: The disease remission rate in the observation group was 62.75%, which was higher than that in the control group (41.18%). After 1 month and 3 months of treatment, the levels of serum CYFRA21-1, CEA, CA125, and IGF-1R were lower, while serum S100A2 was higher in the observation group than in the control group (P < 0.05). For safety assessment, we found that the incidences of neutropenia, thrombocytopenia, and gastrointestinal reactions had no statistical differences between two groups. The time to disease progression in the observation group was 129.85 d longer than that in the control group, 89.74 d, and the six-month survival rate and 1-year survival rate were higher in the observation group relative to the control group. Conclusion: Medical database-based BACE combined with 125I seed implantation in the therapy of advanced NSCLC patients has definite efficacy with certain safety, which can enhance antitumor effect and prolong survival rate in advanced NSCLC patients.

Prognostic Evaluation of CT Imaging Big Data-Assisted Arterial Chemoembolization Combined with 125I Seed Implantation for Non-Small-Cell Lung Cancer.

Objective: To investigate the prognostic impact of computed tomography (CT) imaging big data-assisted arterial chemoembolization combined with iodine 125 (125I) seed implantation on patients with non-small-cell lung cancer (NSCLC). Methods: A total of 116 patients with intermediate and advanced NSCLC hospitalized in our hospital from August 2019 to August 2020 were selected and divided into a control group and an experiment group (58 cases in each group) by random number table method for the study. The patients in the experiment group were treated with CT imaging big data-assisted arterial chemoembolization combined with 125I seed implantation, while the patients in the control group were treated with arterial chemoembolization alone, with the use of gemcitabine combined with cisplatin (GP) in chemotherapy. The prognostic impact was determined by analyzing recent efficacy; the incidence of adverse effects; tumor size and CT perfusion parameters including blood volume (BV), blood flow (BF), and permeability surface (PS); frailty state and quality of life; and the levels of serum tumor markers including carcinoembryonic antigen (CEA), glycoconjugate antigen 125 (CA125), cytokeratin 19 fragment antigen 21-1 (CYFRA21-1), microRNA- (miRNA-) 137, and miR-379-5p. In addition, frailty status was evaluated using the Fried frailty phenotype (FP) scale, and quality of life was determined according to Karnofsky Performance Status (KPS) score. Kaplan-Meyer (KM) method was used to analyze the survival rate of NSCLC patients after a 12-month follow-up. Results: The remission rate in the experiment group (77.59%) was higher than that in the control group (56.90%) (P < 0.05). Tumor size, BV, BF, PS, serum CEA and CA125 levels, and FP value in both groups were dramatically reduced after treatment compared with before treatment, especially in the experiment group after 1 and 3 months of treatment (P < 0.05). Meanwhile, the serum miR-137 and miR-379-5p levels and KPS scores in both groups were higher after treatment than before treatment, especially in the experiment group after 1 and 3 months of treatment (P < 0.05). However, there was no significant difference in the incidence of nausea and vomiting, alopecia, diarrhea, myelosuppression, and hemoptysis of NSCLC patients in both groups after treatment (P > 0.05). Further, the 12-month survival rate of NSCLC patients was higher in the experiment group (84.21%) than in the control group (64.29%) (P < 0.05). Conclusion: CT imaging big data-assisted arterial chemoembolization combined with 125I seed implantation for NSCLC can improve recent efficacy and the prognosis of NSCLC patients by inhibiting tumor progression with a certain degree of safety.

Immune Complex Formation Is Associated With Loss of Tolerance and an Antibody Response to Both Drug and Target.

AMG 966 is a bi-specific, heteroimmunoglobulin molecule that binds both tumor necrosis factor alpha (TNFα) and TNF-like ligand 1A (TL1A). In a first-in-human clinical study in healthy volunteers, AMG 966 elicited anti-drug antibodies (ADA) in 53 of 54 subjects (98.1%), despite a paucity of T cell epitopes observed in T cell assays. ADA were neutralizing and bound to all domains of AMG 966. Development of ADA correlated with loss of exposure. In vitro studies demonstrated that at certain drug-to-target ratios, AMG 966 forms large immune complexes with TNFα and TL1A, partially restoring the ability of the aglycosylated Fc domain to bind FcγRIa and FcγRIIa, leading to the formation of ADA. In addition to ADA against AMG 966, antibodies to endogenous TNFα were also detected in the sera of subjects dosed with AMG 966. This suggests that the formation of immune complexes between a therapeutic and target can cause loss of tolerance and elicit an antibody response against the target.

Dual near-infrared fluorescence-based lateral flow immunosensor for the detection of zearalenone and deoxynivalenol in maize.

A novel dual near-infrared fluorescence-based lateral flow immunosensor was developed to determine zearalenone and deoxynivalenol in maize. Two near-infrared dyes with distinct fluorescence characteristics were utilized to separately label the anti-zearalenone and anti-deoxynivalenol antibodies as detection reagents. The capture antigens zearalenone-BSA and deoxynivalenol-BSA were mixed and immobilized on the same test line of nitrocellulose membrane. This assay format facilitates simultaneous detection of the two mycotoxins on a single test line. After optimizing experimental parameters, the limits of detection for zearalenone and deoxynivalenol were as low as 0.55 µg/kg and 3.8 µg/kg in maize, respectively. The spiking experiment yielded recovery ratios ranging from 81.7% to 107.3% with coefficients of variation less than 14% demonstrating high assay accuracy and precision. Moreover, the actual sample analysis produced consistent results between this method and instrumental method. Therefore, the developed immunosensor can serve as an accurate and efficient approach for monitoring mycotoxins in agricultural products.

Duplex Surface Enhanced Raman Scattering-Based Lateral Flow Immunosensor for the Low-Level Detection of Antibiotic Residues in Milk.

A duplex surface enhanced Raman scattering (SERS)-based lateral flow immunosensor was established for the simultaneous detection of two common antibiotic residues including tetracycline and penicillin in milk. The newly synthesized Au@Ag nanoparticles were labeled with different Raman molecules including 5,5-dithiobis-2-nitrobenzoic acid (DTNB) or 4-mercaptobenzoic acid (MBA), followed by the conjugation of anti-tetracycline monoclonal antibody or anti-penicillin receptor, forming two kinds of SERS nanoprobes. The two nanoprobes can recognize tetracycline-BSA and ampicillin-BSA, respectively, which facilitates the simultaneous detection of the two types of antibiotics on a single test line. After optimization, detection limits of tetracycline and penicillin as low as 0.015 ng/mL and 0.010 ng/mL, respectively, were achieved. These values were far below those of most of other documented bio-analytical approaches. Moreover, the spiking test demonstrates an excellent assay accuracy with recoveries of 88.8% to 111.3%, and satisfactory assay precision with relative standard deviation below 16%. Consequently, the results demonstrate that the SERS-based lateral flow immunosensor developed in this study has the advantages of excellent assay sensitivity and remarkable multiplexing capability, thus it will have great application potential in food safety monitoring.

Interlaboratory Study for Characterizing Monoclonal Antibodies by Top-Down and Middle-Down Mass Spectrometry.

The Consortium for Top-Down Proteomics (www.topdownproteomics.org) launched the present study to assess the current state of top-down mass spectrometry (TD MS) and middle-down mass spectrometry (MD MS) for characterizing monoclonal antibody (mAb) primary structures, including their modifications. To meet the needs of the rapidly growing therapeutic antibody market, it is important to develop analytical strategies to characterize the heterogeneity of a therapeutic product's primary structure accurately and reproducibly. The major objective of the present study is to determine whether current TD/MD MS technologies and protocols can add value to the more commonly employed bottom-up (BU) approaches with regard to confirming protein integrity, sequencing variable domains, avoiding artifacts, and revealing modifications and their locations. We also aim to gather information on the common TD/MD MS methods and practices in the field. A panel of three mAbs was selected and centrally provided to 20 laboratories worldwide for the analysis: Sigma mAb standard (SiLuLite), NIST mAb standard, and the therapeutic mAb Herceptin (trastuzumab). Various MS instrument platforms and ion dissociation techniques were employed. The present study confirms that TD/MD MS tools are available in laboratories worldwide and provide complementary information to the BU approach that can be crucial for comprehensive mAb characterization. The current limitations, as well as possible solutions to overcome them, are also outlined. A primary limitation revealed by the results of the present study is that the expert knowledge in both experiment and data analysis is indispensable to practice TD/MD MS.

Surface enhanced Raman scattering-based lateral flow immunosensor for sensitive detection of aflatoxin M1 in urine.

Aflatoxin M1 (AFM1) is generally used as a biomarker in urine for the assessment of aflatoxin exposure in humans and animals. However, there is no approach for the rapid and on-site monitoring of AFM1 level in urine. Here, we report a surface enhanced Raman scattering (SERS)-based lateral flow immunosensor built for such a purpose. Raman molecule 5,5-dithiobis-2-nitrobenzoic acid and anti-AFM1 monoclonal antibody were conjugated with Au (core)@Ag (shell) nanoparticle to serve as SERS nanoprobe. AFM1-bovine serum albumin was conjugated with gold nanoparticle and then applied onto nitrocellulose membrane as a visible "GOLD" test line. Quantitation of AFM1 was performed by the readout of Raman signal from the SERS nanoprobes captured on the test line. After optimizing experimental parameters, the detection limit of this immunosensor can achieve as low as 1.7 pg/mL of AFM1 in urine, which is far below the recommended tolerable level (30 pg/mL) of AFM1 in urine. The spiking experiment yielded 93.8%-111.3% recovery with coefficients of variation below 17% demonstrating high assay accuracy and precision. Moreover, this immunosensing assay is fast with an assay time below 20 min. Therefore, the developed immunosensor is a promising tool for the rapid assessment of aflatoxin exposure in the field.

Lateral flow immunoassay for 5-hydroxyflunixin based on near-infrared fluorescence molecule as an alternative label to gold nanoparticles.

A high-affinity monoclonal antibody (mAb) has been prepared and separately a gold nanoparticle (AuNP)-based and a near-infrared (NIR) fluorescence-based lateral flow immunoassay (LFA) developed for determination of 5-hydroxyflunixin residue in raw milk. The AuNP and IRDye® 800CW were used to label anti-5-hydroxyflunixin mAb to form the AuNP-mAb and NIR dye-mAb conjugates, respectively. Quantitative determination of 5-hydroxyflunixin was achieved by imaging the optical or fluorescence intensity of the AuNP-mAb and NIR dye-mAb captured on the test line. As a result, the detection limits of the AuNP-based LFA and NIR dye-based LFA were 0.82 and 0.073 ng/mL in raw milk, respectively. The considerable improvement on assay sensitivity of the NIR-based LFA can be attributed to the lower background and less antibody consumption per test than that of the AuNP-based LFA. The spiking experiment by the NIR-based LFA yielded 85.7-112.6% recovery with a relative standard deviation below 14%, indicating that it has satisfactory assay accuracy and precision. Furthermore, the analytical results of actual samples by the NIR dye-based LFA were consistent with that by instrumental analysis. Therefore, these results demonstrated that the NIR dye is an ideal alternative label to the conventional AuNP for the development of LFA for veterinary drugs in animal-origin food. Graphical abstract.

Multiplex SERS-based lateral flow immunosensor for the detection of major mycotoxins in maize utilizing dual Raman labels and triple test lines.

A multiplex surface-enhanced Raman scattering (SERS)-based lateral flow immunosensor was developed to determine six major mycotoxins in maize. Two characteristic Raman reporter molecules-5,5-dithiobis-2-nitrobenzoic acid (DTNB) and 4-mercaptobenzoic acid (MBA)-were used to label the synthesized Au@Ag core-shell nanoparticles for the preparation of SERS nanoprobes as detection reagents. Six corresponding hapten-protein conjugates were prepared and dispensed on three test lines of nitrocellulose membrane with two conjugates on each line as capture antigens. This design facilitates the simultaneous detection of the six mycotoxins in a single test. After optimizing the experimental parameters of immunosensor, the limits of detection were as low as 0.96 pg/mL for aflatoxin B1, 6.2 pg/mL for zearalenone, 0.26 ng/mL for fumonisin B1, 0.11 ng/mL for deoxynivalenol, 15.7 pg/mL for ochratoxin A, and 8.6 pg/mL for T-2 toxin, respectively. The spiking experiment showed high accuracy with recovery of 78.9-106.2 % and satisfactory assay precision with the coefficient of variations below 16 %. Moreover, this assay can be completed in less than 20 min, and its detection results were consistent with that of liquid chromatography-mass spectrometry. Therefore, the developed SERS-based lateral flow immunosensor is a promising approach for mycotoxin detection in the field.

Rapid one-step enzyme immunoassay and lateral flow immunochromatographic assay for colistin in animal feed and food.

BACKGROUND: Colistin (polymyxin E) is a kind of peptide antibiotic which has been approved in animal production for the purposes of disease prevention, treatment, and growth promotion. However, the wide use of colistin in animal feed may accelerate the spread of colistin-resistance gene MCR-1 from animal production to human beings, and its residue in animal-origin food may also pose serious health hazards to humans. Thus, it is necessary to develop corresponding analytical methods to monitor the addition of colistin in animal feed and the colistin residue in animal-origin food. RESULTS: A one-step enzyme-linked immunosorbent assay (ELISA) and a lateral flow immunochromatographic assay (LFIA) for colistin were developed based on a newly developed monoclonal antibody. The ELISA showed a 50% inhibition value (IC50) of 9.7 ng/mL with assay time less than 60 min, while the LFIA had a strip reader-based detection limit of 0.87 ng/mL in phosphate buffer with assay time less than 15 min. For reducing the non-specific adsorption of colistin onto sample vial, the components of sample extraction solution were optimized and proved to greatly improve the assay accuracy. The spiked recovery experiment showed that the recoveries of colistin from feed, milk and meat samples were in the range of 77.83% to 113.38% with coefficient of variations less than 13% by ELISA analysis and less than 18% by LFIA analysis, respectively. Furthermore, actual sample analysis indicated that the two immunoassays can produce results consistent with instrumental analysis. CONCLUSIONS: The developed assays can be used for rapid qualitative or quantitative detection of colistin in animal feed and food.

Quantitative detection and evaluation of thrombus formation based on electrical impedance spectroscopy.

Thrombus formation is quantitatively measured and evaluated by the electrical impedance spectroscopy method in this study, which confirms the possibility for the application of a promising non-invasive thrombus detection method. The impedance parameter Z*(t) of blood from the electrical impedance spectroscopy is utilized to elaborate the impedance performance of blood during thrombus formation process. Experimental results indicate that the impedance Z*(t) of blood has regular variations under the formation of thrombus, which could be divided into three stages. Modified Hanai equation is proposed to quantitatively expound the three stages of impedance Z*(t) variation. The amount of fibrin and thrombus clot is founded to be accounted for the impedance variation of blood, which confirms the feasibility and theoretical basis of the non-invasive and on-line thrombus bio-detection technology for patients with serious cardiovascular disease.

Diagnosis of Hemoglobinopathy and ß-Thalassemia by 21 Tesla Fourier Transform Ion Cyclotron Resonance Mass Spectrometry and Tandem Mass Spectrometry of Hemoglobin from Blood.

BACKGROUND: Hemoglobinopathies and thalassemias are the most common genetically determined disorders. Current screening methods include cation-exchange HPLC and electrophoresis, the results of which can be ambiguous because of limited resolving power. Subsequently, laborious genetic testing is required for confirmation. METHODS: We performed a top-down tandem mass spectrometry (MS/MS) approach with a fast data acquisition (3 min), ultrahigh mass accuracy, and extensive residue cleavage by use of positive electrospray ionization 21 Tesla Fourier transform ion cyclotron resonance-tandem mass spectrometry (21 T FT-ICR MS/MS) for hemoglobin (Hb) variant de novo sequencing and ß-thalassemia diagnosis. RESULTS: We correctly identified all Hb variants in blind analysis of 18 samples, including the first characterization of homozygous Hb Himeji variant. In addition, an Hb heterozygous variant with isotopologue mass spacing as small as 0.0194 Da (Hb AD) was resolved in both precursor ion mass spectrum (MS1) and product ion mass spectrum (MS2). In blind analysis, we also observed that the abundance ratio between intact δ and ß subunits (δ/ß) or the abundance ratio between intact δ and α subunits (δ/α) could serve to diagnose ß-thalassemia trait caused by a mutation in 1 HBB gene. CONCLUSIONS: We found that 21 T FT-ICR MS/MS provides a benchmark for top-down MS/MS analysis of blood Hb. The present method has the potential to be translated to lower resolving power mass spectrometers (lower field FT-ICR mass spectrometry and Orbitrap) for Hb variant analysis (by MS1 and MS2) and ß-thalassemia diagnosis (MS1).

Classification of Plasma Cell Disorders by 21 Tesla Fourier Transform Ion Cyclotron Resonance Top-Down and Middle-Down MS/MS Analysis of Monoclonal Immunoglobulin Light Chains in Human Serum.

The current five-year survival rate for systemic AL amyloidosis or multiple myeloma is ∼51%, indicating the urgent need for better diagnosis methods and treatment plans. Here, we describe highly specific and sensitive top-down and middle-down MS/MS methods owning the advantages of fast sample preparation, ultrahigh mass accuracy, and extensive residue cleavages with 21 telsa FT-ICR MS/MS. Unlike genomic testing, which requires bone marrow aspiration and may fail to identify all monoclonal immunoglobulins produced by the body, the present method requires only a blood draw. In addition, circulating monoclonal immunoglobulins spanning the entire population are analyzed and reflect the selection of germline sequence by B cells. The monoclonal immunoglobulin light chain FR2-CDR2-FR3 was sequenced by database-aided de novo MS/MS and 100% matched the gene sequencing result, except for two amino acids with isomeric counterparts, enabling accurate germline sequence classification. The monoclonal immunoglobulin heavy chains were also classified into specific germline sequences based on the present method. This work represents the first application of top/middle-down MS/MS sequencing of endogenous human monoclonal immunoglobulins with polyclonal immunoglobulins background.

Quadruplex gold immunochromatogaraphic assay for four families of antibiotic residues in milk.

In this study, we developed a quadruplex gold immunochromatogaraphic assay (GICA) for the simultaneous determination of four families of antibiotics including ß-lactams, tetracyclines, streptomycin and chloramphenicol in milk. For qualitative analysis, the visual cut-off values were measured to be 2-100 ng/mL, 16-32 ng/mL, 50 ng/mL and 2.4 ng/mL for ß-lactams, tetracyclines, streptomycin and chloramphenicol, respectively. For quantitative analysis, the detection ranges were 0.13-1 ng/mL for penicillin G, 0.13-8 ng/mL for tetracycline, 0.78-25 ng/mL for streptomycin, 0.019-1.2 ng/mL for chloramphenicol in milk respectively, with linear correlation coefficients higher than 0.97. The spiked experiment indicated that the mean recoveries ranged from 84.5% to 107.6% with coefficient of variations less than 16.2%, and real sample analysis revealed that the GICA can produce consistent results with instrumental analysis. These results demonstrated that this novel immunoassay is a promising approach for rapidly screening common antibiotic residues in milk.

Accurate Identification of Unknown and Known Metabolic Mixture Components by Combining 3D NMR with Fourier Transform Ion Cyclotron Resonance Tandem Mass Spectrometry.

Metabolite identification in metabolomics samples is a key step that critically impacts downstream analysis. We recently introduced the SUMMIT NMR/mass spectrometry (MS) hybrid approach for the identification of the molecular structure of unknown metabolites based on the combination of NMR, MS, and combinatorial cheminformatics. Here, we demonstrate the feasibility of the approach for an untargeted analysis of both a model mixture and E. coli cell lysate based on 2D/3D NMR experiments in combination with Fourier transform ion cyclotron resonance MS and MS/MS data. For 19 of the 25 model metabolites, SUMMIT yielded complete structures that matched those in the mixture independent of database information. Of those, seven top-ranked structures matched those in the mixture, and four of those were further validated by positive ion MS/MS. For five metabolites, not part of the 19 metabolites, correct molecular structural motifs could be identified. For E. coli, SUMMIT MS/NMR identified 20 previously known metabolites with three or more 1H spins independent of database information. Moreover, for 15 unknown metabolites, molecular structural fragments were determined consistent with their spin systems and chemical shifts. By providing structural information for entire metabolites or molecular fragments, SUMMIT MS/NMR greatly assists the targeted or untargeted analysis of complex mixtures of unknown compounds.

Analysis of Monoclonal Antibodies in Human Serum as a Model for Clinical Monoclonal Gammopathy by Use of 21 Tesla FT-ICR Top-Down and Middle-Down MS/MS.

With the rapid growth of therapeutic monoclonal antibodies (mAbs), stringent quality control is needed to ensure clinical safety and efficacy. Monoclonal antibody primary sequence and post-translational modifications (PTM) are conventionally analyzed with labor-intensive, bottom-up tandem mass spectrometry (MS/MS), which is limited by incomplete peptide sequence coverage and introduction of artifacts during the lengthy analysis procedure. Here, we describe top-down and middle-down approaches with the advantages of fast sample preparation with minimal artifacts, ultrahigh mass accuracy, and extensive residue cleavages by use of 21 tesla FT-ICR MS/MS. The ultrahigh mass accuracy yields an RMS error of 0.2-0.4 ppm for antibody light chain, heavy chain, heavy chain Fc/2, and Fd subunits. The corresponding sequence coverages are 81%, 38%, 72%, and 65% with MS/MS RMS error ~4 ppm. Extension to a monoclonal antibody in human serum as a monoclonal gammopathy model yielded 53% sequence coverage from two nano-LC MS/MS runs. A blind analysis of five therapeutic monoclonal antibodies at clinically relevant concentrations in human serum resulted in correct identification of all five antibodies. Nano-LC 21 T FT-ICR MS/MS provides nonpareil mass resolution, mass accuracy, and sequence coverage for mAbs, and sets a benchmark for MS/MS analysis of multiple mAbs in serum. This is the first time that extensive cleavages for both variable and constant regions have been achieved for mAbs in a human serum background. Graphical Abstract ᅟ.