Application of the cell-based RT-qPCR assay (C-QPA) for potency detection of the novel trivalent rotavirus vaccine in China.

BACKGROUND: Because of the deficiencies of traditional methods in multivalent rotavirus vaccine potency detection, a cell-based quantitative RT-qPCR assay (C-QPA) was established and validated for specificity, precision, and accuracy. METHODS: In order to further validate the robustness of this method in actual titer detection, the linear range and the practical application under different conditions were tested using monovalent and trivalent rotavirus samples and standards. RESULTS: Results showed that the linear range was 2.0-6.5, 3.9-8.3, and 3.5-8.1 UI (unit of infectivity) for G2, G3, and G4, respectively. Besides, unknown sample with high titer exceeding the linear range can be calculated by dilution. The UIs of serotypes G2, G3, and G4 in monovalent and trivalent rotavirus samples showed a relative deviation ≤4.10%, and the monovalent samples of the same serotype with or without protective agents showed a relative deviation ≤4.28%; the coefficient of variation (CV) of at least 176 tests (548 individual runs) of 3 in vitro-transcribed RNA standards with certain concentrations was not higher than 6.50%; the results of the trivalent samples tested by more than 149 times in 5 years (467 individual runs) showed the CVs lower than 12.66%; 15 samples detected by one laboratory showed a CV lower than 9.83%, while other three samples tested by two independent laboratories showed a CV lower than 6.90%. CONCLUSION: In summary, the C-QPA has good linearity, durability, repeatability, and reproducibility in practical application and has been proved by the authority to be widely used in the production, quality control and release of the recently licensed trivalent vaccine in China.

The effect of extracellular polymeric substances (EPS) of iron-oxidizing bacteria (Ochrobactrum EEELCW01) on mineral transformation and arsenic (As) fate.

Extracellular polymeric substances (EPS) are an important medium for communication and material exchange between iron-oxidizing bacteria and the external environment and could induce the iron (oxyhydr) oxides production which reduced arsenic (As) availability. The main component of EPS secreted by iron-oxidizing bacteria (Ochrobactrum EEELCW01) was composed of polysaccharides (150.76-165.33 mg/g DW) followed by considerably smaller amounts of proteins (12.98-16.12 mg/g DW). Low concentrations of As (100 or 500 µmol/L) promoted the amount of EPS secretion. FTIR results showed that EPS was composed of polysaccharides, proteins, and a miniscule amount of nucleic acids. The functional groups including -COOH, -OH, -NH, -C=O, and -C-O played an important role in the adsorption of As. XPS results showed that As was bound to EPS in the form of As3+. With increasing As concentration, the proportion of As3+ adsorbed on EPS increased. Ferrihydrite with a weak crystalline state was only produced in the system at 6 hr during the mineralization process of Ochrobactrum sp. At day 8, the minerals were composed of goethite, galena, and siderite. With the increasing mineralization time, the main mineral phases were transformed from weakly crystalline hydrous iron ore into higher crystallinity siderite (FeCO3) or goethite (α-FeOOH), and the specific surface area and active sites of minerals were reduced. It can be seen from the distribution of As elements that As is preferentially adsorbed on the edges of iron minerals. This study is potential to understand the biomineralization mechanism of iron-oxidizing bacteria and As remediation in the environment.

Elsholtzia ciliata (Thunb.) Hyland: A Review of Phytochemistry and Pharmacology.

In this paper, the confusion of the sources of medicinal materials was briefly expounded, and the differences among the varieties were pointed out. At the same time, the chemical components and pharmacological properties of Elsholtzia ciliata (Thunb.) Hyland (E. ciliata) were reviewed. The structures of 352 compounds that have been identified are listed. These mainly include flavonoids, terpenoids, phenylpropanoids, alkaloids, and other chemical components. They have antioxidant, anti-inflammatory, antimicrobial, insecticidal, antiviral, hypolipidemic, hypoglycemic, analgesic, antiarrhythmic, antitumor, antiacetylcholinesterase, and immunoregulator activities. At present, there are many researches using essential oil and alcohol extract, and the researches on antioxidant, anti-inflammatory, anti-microbial, and other pharmacological activities are relatively mature. This paper aims to summarize the existing research, update the research progress regarding the phytochemicals and pharmacology of E. ciliate, and to provide convenience for subsequent research.

The Genus Broussonetia: An Updated Review of Phytochemistry, Pharmacology and Applications.

The Broussonetia genus (Moraceae), recognized for its value in many Chinese traditional herbs, mainly includes Broussonetia papyrifera (L.) L'Hér. ex Vent. (BP), Broussonetia kazinoki Siebold (BK), and Broussonetia luzonica (Blanco) Bureau (BL). Hitherto, researchers have found 338 compounds isolated from BP, BK, and BL, which included flavonoids, polyphenols, phenylpropanoids, alkaloids, terpenoids, steroids, and others. Moreover, its active compounds and extracts have exhibited a variety of pharmacological effects such as antitumor, antioxidant, anti-inflammatory, antidiabetic, anti-obesity, antibacterial, and antiviral properties, and its use against skin wrinkles. In this review, the phytochemistry and pharmacology of Broussonetia are updated systematically, after its applications are first summarized. In addition, this review also discusses the limitations of investigations and the potential direction of Broussonetia. This review can help to further understand the phytochemistry, pharmacology, and other applications of Broussonetia, which paves the way for future research.

Cynanchum auriculatum Royle ex Wight., Cynanchum bungei Decne. and Cynanchum wilfordii (Maxim.) Hemsl.: Current Research and Prospects.

Cynanchum auriculatum Royle ex Wight. (CA), Cynanchum bungei Decne. (CB) and Cynanchum wilfordii (Maxim.) Hemsl. (CW) are three close species belonging to the Asclepiadaceous family, and their dry roots as the bioactive part have been revealed to exhibit anti-tumor, neuroprotection, organ protection, reducing liver lipid and blood lipid, immunomodulatory, anti-inflammatory, and other activities. Until 2021, phytochemistry investigations have uncovered 232 compounds isolated from three species, which could be classified into C21-steroids, acetophenones, terpenoids, and alkaloids. In this review, the morphology characteristics, species identification, and the relationship of botany, extraction, and the separation of chemical constituents, along with the molecular mechanism and pharmacokinetics of bioactive constituents of three species, are summarized for the first time, and their phytochemistry, pharmacology, and clinical safety are also updated. Moreover, the direction and limitation of current research on three species is also discussed.

Establishment of a cell-based quantitative reverse transcription-polymerase chain reaction (RT-qPCR) assay for detection of multivalent rotavirus vaccine.

Because of deficiencies of traditional potency tests in rotavirus detection, a one-step TaqMan probe-based quantitative reverse transcription-polymerase chain reaction (RT-qPCR) assay combined with cell-based method was established to determine the infectious potency of the target virus in multivalent live rotavirus vaccines in vitro. Series dilutions of rotavirus samples were inoculated into Vero cells and cultured for 24 hours. The cells were lysed and the potency was detected by RT-qPCR. The reference standards with a known titer (lgCCID50 /mL) were assayed in parallel, and the potencies of each sample were determined using parallel line method. The specificity, precision and accuracy of the assay were evaluated, respectively. The results showed that messenger RNA produced during rotavirus replication was the primary template of RT-qPCR and the primers and probes were specific to each strain. The coefficient of variation of different wells and different working days did not exceed 6% and the results of the assay were proved to be concordant with those of cell culture infective dose 50% with a relative deviation less than 5%. This assay is a more rapid, cost-effective and high-throughput way for detecting multivalent rotavirus vaccine, and will be a valuable tool in the quality control and stability monitoring of live multivalent rotavirus vaccine.

Achyranthes bidentata Blume (Amaranthaceae): a review of its botany, traditional uses, phytochemistry, pharmacology, and toxicology.

OBJECTIVES: Achyranthes bidentata Blume (A. bidentata) is a plant of Amaranthaceae family, and its root is the main medicinal part, named "Huai-Niu-Xi." It is used to expel blood stasis through menstruation, tonify liver and kidney, strengthen muscles and bones, and induce diuresis. This review aimed to provide a systematic summary of botany, traditional uses, phytochemistry, pharmacology, and toxicology of A. bidentata. METHODS: The present review covers the literature survey. The data have been collected from various journals, books, and some of the electronic search via Internet-based information such as Web of Science, PubMed, Google Scholar, Google patents, CNKI, SpringerLink, online electronic journals, and ScienceDirect. KEY FINDINGS: So far, more than 270 metabolites have been isolated from A. bidentata, including terpenoids, steroids, alkaloids, flavonoids, and so on. Among them, terpenoids and steroids are the main metabolites. The extract and metabolites exert multiple pharmacological activities such as alleviating osteoarthritis effect, antiosteoporosis activity, neuroprotective effect, antidiabetic activity-associated complications, immunoregulatory activity, and so on. SUMMARY: Some traditional uses of A. bidentata need further in-depth studies to confirm. Similarly, the separation and screening of active compounds, as well as the corresponding molecular mechanisms of action of compounds, are also needed to be studied.

[Research Progress on Solidification and MICP Remediation of Soils in Heavy Metal Contaminated Site].

Heavy metal pollution in soils of smelting sites is an important environmental problem to be solved urgently. Solidification technology has become one of the mainstream technologies for heavy metal remediation in contaminated sites owing to its shorter remediation time, low cost, and high treatment efficiency. On the basis of summarizing the latest research progress on the remediation of heavy metal pollution in sites by solidification in the past 10 years, this study focused on the mechanisms of solidification technology and analyzed the advantages and disadvantages of different mechanisms (mechanism of inorganic materials, mechanism of organic materials, mechanism of mechanical ball milling, and mechanism of microbial-induced carbonate mineralization (MICP)) and their scope of application. Then, according to the research focus and development trend presented by CiteSpace, the application prospects and limiting factors of MICP technology for the solidification and remediation of heavy metal pollution in sites were summarized from three aspects:the application of MICP in multi-metal remediation, the application of MICP composites in contaminated sites, and the influencing factors of MICP technology application. Finally, the prospects and challenges in solidification technology were put forward in order to provide reference for the future development.

The genus Oxytropis DC: application, phytochemistry, pharmacology, and toxicity.

OBJECTIVES: Oxytropis DC is a perennial plant of Fabaceae family, which is widely distributed in the northern temperate zone. It is known as "locoweed" because of its toxic component swainsonine. However, it is widely used in Tibetan medicine and Mongolian medicine, mainly for the treatment of heat-clearing and detoxifying, pain-relieving, anti-inflammatory, hemostasis, and other diseases. To provide a basis for the further development and utilization of Oxytropis DC, the pieces of literature about the application, phytochemistry, pharmacological action, and toxicity of Oxytropis DC were reviewed and analyzed. KEY FINDINGS: A total of 373 chemical constituents were found from Oxytropis DC, including flavonoids, alkaloids, steroids, terpenoids, and others. Pharmacological actions mainly include antitumor, antioxidation, anti-inflammatory, analgesic, antibacterial, antifibrosis, and other pharmacological actions, among them, the antitumor effect is particularly prominent. SUMMARY: At present, studies on its pharmacological effects are mainly concentrated on the extracts, some flavonoids, and alkaloids. In the follow-up studies, research on the pharmacological activities of the other chemical constituents in Oxytropis should be strengthened. It has the potential to pave the way for research and development of novel Oxytropis medicines.

Ligusticum chuanxiong Hort.: a review of its phytochemistry, pharmacology, and toxicology.

BACKGROUND: Conioselinum anthriscoides (H. Boissieu) Pimenov & Kljuykov, also known as Ligusticum chuanxiong Hort. is a perennial Umbelliferae herb, whose dried rhizome commonly called Chuanxiong Rhizoma. Chuanxiong Rhizoma is widely used in TCM, especially for cardiocerebrovascular and gynecological diseases. However, these studies are scattered and there is no review that can centralize the results of these studies. The authors summarized this review by collecting research results on the chemical, pharmacological, and toxicological of Chuanxiong Rhizoma published in various publications over the past 20 years. AIMS: The purpose of this review is to summarize the current experimental studies on Chuanxiong Rhizoma and explore its mechanism of action. METHODS: Web of Science, PubMed, CBM, CNKI, Medline, Embase, Elsevier, Springer, Wiley Online Library, Scholar, and other databases were searched, and nearly one hundred experimental studies were collected to summarize this review. RESULTS AND DISCUSSION: Chuanxiong Rhizoma is composed of essential oil, terpenes, alkaloids, polysaccharide, organic acids, ceramides, and cerebrosides. It has the functions of promoting blood circulation, removing blood stasis, antibacterial, antiviral, and calming the mind to sleep. Now it can be used to treat cardiocerebrovascular and gynecological diseases, neurodegenerative disease, psoriasis, rectal cancer, osteoporosis, and osteoarthritis. CONCLUSIONS: In the past 20 years, a large number of research data have confirmed that Chuanxiong Rhizoma contains rich effective metabolites, has huge medicinal potential, and has a wide range of effective treatments.

Polysaccharide from Panax japonicus C.A. Mey prevents non-alcoholic fatty liver disease development based on regulating liver metabolism and gut microbiota in mice.

In this study, a new polysaccharide (PSPJ) with specific molecular weight and monosaccharide compositions was isolated and purified from the water extract of Panacis Japonici Rhizoma (PJR). 16S rRNA analysis and untargeted metabolomic analysis were used to assess PSPJ's efficacy in averting non-alcoholic fatty liver disease (NAFLD). This study indicated that PSPJ significantly reduced liver fat accumulation, the increase in blood lipids and ALT caused by HFD, indicating that PSPJ can prevent NAFLD. We demonstrated through cell experiments that PSPJ does not directly affect liver cells. The gut microbiota disorder and alterations in short-chain fatty acids (SCFAs) induced by the high-fat diet (HFD) were ameliorated by PSPJ, as evidenced by the analysis of 16S rRNA. In particular, supplementing PSPJ reduced the abundance of Turicibacter, Dubosiella, and Staphylococcus, and increased the abundance of Bacteroides, Blautia, and Lactobacillus. Untargeted metabolomic analysis shows that PSPJ improves liver metabolic disorders by regulating arachidonic acid metabolism, carbohydrate digestion and absorption, fatty acid biosynthesis, fatty acid metabolism and retinol metabolism. The findings of our investigation indicate that PSPJ has the potential to modulate liver metabolism through alterations in the composition of intestinal bacteria, hence preventing NAFLD.

Cardioprotective effect of crude polysaccharide fermented by Trametes Sanguinea Lyoyd on doxorubicin-induced myocardial injury mice.

Doxorubicin (DOX) is a broad-spectrum anti-tumor drug, but its clinical application is greatly limited because of the cardiotoxicity. Thus, exploration of effective therapies against DOX-induced cardiotoxicity is necessary. The aim of this study is to investigate the effects and possible mechanisms of Trametes Sanguinea Lyoyd fermented crude polysaccharide (TSLFACP) against DOX-induced cardiotoxicity. We investigated the protective effects of TSLFACP on myocardial injury and its possible mechanisms using two in vitro cells of DOX-treated cardiomyocytes H9C2 and embryonic myocardial cell line CCC-HEH-2 and a in vivo mouse model of DOX-induced myocardial injury. We found that TSLFACP could reverse DOX-induced toxicity in H9C2 and CCC-HEH-2 cells. Similarly, we found that when pretreatment with TSLFACP (200 mg/kg, i.g.) daily for 6 days, DOX-induced myocardial damage was attenuated, including the decrease in serum myocardial injury index, and the amelioration in cardiac histopathological morphology. Additionally, immunohistochemistry and western blotting were used to identify the underlying and possible signal pathways. We found that TSLFACP attenuated the expression of LC3-II, Beclin-1 and PRAP induced by DOX. In conclusion, our results demonstrated that TSLFACP could protect against DOX-induced cardiotoxicity by inhibiting autophagy and apoptosis.

Dendrophthoe falcata (L.f.) Ettingsh. and Dendrophthoe pentandra (L.) Miq.: A review of traditional medical uses, phytochemistry, pharmacology, toxicity, and applications.

Dendrophthoe falcata (L.f.) Ettingsh. (DF) and Dendrophthoe pentandra (L.) Miq. (DP) have been traditionally used for the treatment of various ailments, such as cancer, ulcers, asthma, paralysis, skin diseases, tuberculosis, and menstrual troubles, in the ethnomedicinal systems of India and Indonesia. Currently, the chemical structures of 46 compounds have been elucidated from DF and DP, including flavonoids, triterpenes, tannins, steroids, open-chain aliphatics, benzyl derivates, and cyclic chain derivatives. In vitro assays have revealed their anti-tumor and anti-microbial activities. In vivo studies have unraveled their pharmacological properties against tumors, depression, fertility disorders, inflammatory responses, and so on. Additionally, their weak toxicity to rats and brine shrimp, as well as their promising applications for pharmaceutical preparations and combined medication, were also revealed. Herein, we not only recapitulated traditional medical uses, phytochemistry, pharmacology, toxicity, and applications of DF and DP but also discussed current research limitations and future perspectives, which are instructive for those interested in them and are committed to advancing parasitic plants to the Frontier of phytomedicine. We highlighted that DF and DP will become promising medical plants rather than being discarded as notorious pests, provided that more and deeper research is undertaken.

Research progress on chemical components and pharmacological action of Solanum lyratum Thunb.

OBJECTIVES: Solanum lyratum Thunb (SLT) is a perennial plant of the Solanaceae family, and is extensively used in the clinical practice of traditional Chinese medicine. Malaria, oedema, gonorrhoea, cancer, wind and fever, jaundiced hepatitis, cholecystitis and rheumatoid arthritis are among the diseases that it is used to treat. To offer a foundation for further development and usage of SLT, the pieces of literature about the chemical composition and pharmacological action of SLT were reviewed and analysed. KEY FINDINGS: The chemical constituents of SLT mainly included steroids, alkaloids, flavonoids, terpenoids, anthraquinones, phenylpropanoids and others. Pharmacological action mainly contains anti-tumour, antibacterial, anti-inflammatory, anti-oxidation and other pharmacological actions, among them, the anti-tumour effect is particularly outstanding. SUMMARY: At present, studies on the pharmacological effects of SLT mainly focus on alkaloids and steroidal saponins. In the follow-up studies, studies on the pharmacological activities of other chemical components in SLT, such as flavonoids and terpenoids, should be strengthened. It has the potential to pave the way for more research and development of novel SLT medicines.

Tapinanthus species: A review of botany and biology, secondary metabolites, ethnomedical uses, current pharmacology and toxicology.

ETHNOPHARMACOLOGICAL RELEVANCE: Tapinanthus species are hemiparasites that grow on diverse hosts in African regions. Tapinanthus species are locally known as "all purpose herbs" as they are traditionally used to treat various diseases such as diabetes, hypertension, cancer, inflammation, malaria, anemia, anxiety, itching, and so on. AIM OF THE STUDY: A comprehensive review on research outcomes and future perspectives of Tapinanthus species are presented to provide a reference for relevant researchers. MATERIALS AND METHODS: The references regarding Tapinanthus species were retrieved from Google Scholar, Web of Science, Sci-finder, PubMed, Elsevier, Wiley, China National Knowledge Infrastructure, Open Access Library, and SpringerLink between 1963 and 2022. Scientific plant names were provided by "The Plant List" (www.theplantlist.org) and "The world Flora Online" (www.worldfloraonline.org). RESULTS: Even though Tapinanthus species are regarded as notorious pests that can undermine various hosts, they are, as omnipotent herbs in folklore, meaningful for the development of potential phytomedicine sources. Phytochemistry screening has revealed the presence of glycosides, triterpenoids, flavonoids, alkaloids, tannins, steroids, anthraquinones. Among them, the chemical structures of 40 compounds have been elucidated by phytochemical methods without alkaloids and anthraquinones. These secondary metabolites might be responsible for ethnomedical uses and bioactivities of Tapinanthus species. Current research has provided scientific evidence for traditional uses of Tapinanthus species, especially unraveling hypoglycemic, hepatoprotective, antioxidant, antibacterial, anti-anxiety, anti-depression, anti-inflammatory, and other pharmacological properties. Given the fact that ethnomedical uses served as a valuable reference for pharmacology, however, some records to treat arthritis, fever, itching, dysentery, stomach pain, and anemia, have not been confirmed in current research. Furthermore, the toxic effects of Tapinanthus species were susceptible to the dosages, with relative safety across a wide range. CONCLUSIONS: To reasonably yield Tapinanthus species, artificial culture might be a promising method to develop in the future. The discrepancies between phytochemistry screening and structure elucidation, as well as between ethnomedical uses and current pharmacology, need to be further clarified. The identification of bioactive compounds in crude extracts and fractions, the illustration of the underlying mechanisms of pharmacology, along with the addition of cytotoxicity, genotoxicity, and clinical trials of toxic tests, should be carried out in depth. This review highlights that Tapinanthus species can be considered promising phytomedicine sources as long as we adhere to digging more deeply into their potential role.

Design, synthesis, and evaluation of 9-(pyrimidin-2-yl)-9H-carbazole derivatives disrupting mitochondrial homeostasis in human lung adenocarcinoma.

Since more than 85% of lung cancer cases are non-small cell lung cancer (NSCLC), finding novel agents with anti-tumor activities is meaningful for NSCLC patients. Mitochondria is essential for cellular energy metabolism in cancer, and regulating mitochondrial bioenergetics is emerging as a practical approach for cancer treatment and prevention. The carbazole scaffold is an active structure showing anti-cancer biological activity, and the structural diversity has been expanded through the improvement and optimization of synthesizing methods. To find novel carbazole derivatives with great anti-tumor potential and explore structures variety, we designed and synthesized a series of 9-(pyrimidin-2-yl)-9H-carbazole derivatives based on the previously reported Cp∗Rh(III)/H+ tandem catalytic system. With thoroughly bioactivity exploration, we found benzo[d] [1,3]dioxol-5-yl(9-(pyrimidin-2-yl)-9H-carbazol-1-yl)methanone (compound 5n) showed notable activity in disrupting the mitochondrial homeostasis, induced cell cycle arrest and apoptosis in human adenocarcinoma cells, and finally showed anti-tumor activity in an NSCLC-xenograft mice model.

The Improvement and Application of Lentivirus-Mediated Gene Transfer and Expression System in Penaeid Shrimp Cells.

This study first reported the improvement and application of lentivirus-mediated gene transfer and expression system in shrimp cells. After modified by the inclusion of two envelope proteins (VP19 and VP28) of shrimp white spot syndrome virus (WSSV) into the envelope of the packaged lentivirus, and insertion of a truncated promoter of immediate-early gene 1 (Pie1-504) of shrimp WSSV virus into the lentiviral reporter plasmid, the second-generation lentiviral expression system (pLVX-PEF1α-IRES-mCherry, psPAX2, and PMD2.G) was found to behave better in the mitosis-arrested shrimp cells than the similarly modified retrovirus expression system did. Results from the insect sf9 cells indicated that the inclusion of VP19 and VP28 into the envelope of packaged lentiviruses could significantly improve the tropism or infectivity of the modified lentiviruses to insect cells in a cumulative way. Notably, the VP28 contributed about 86% of the total increase of the tropism. In the shrimp primary lymphoid cells infected by modified lentivirus IV with both VP19 and VP28 included, the infection efficiency was up to 11% (non-confocal) and 19% (confocal) and no background fluorescent signal was observed. However, background fluorescent signal was observed in the shrimp primary Oka organ cells although only under a confocal microscope. In the lentivirus IV-infected Oka organ cells, the actual infection efficiencies were calculated up to 8% (non-confocal) and 19% (confocal), significantly higher than those of commercial intact lentivirus I of 0 (non-confocal) and 3% (confocal). The insertion of WSSV promoter (Pie1-504) had interrupted the effective expression of reporter plasmid encoding lentiviral construct of pLVX-PEF1α-Pie1-504-IRES-mCherry in the HEK293T cells, but markedly increased its efficiencies up to 14% (non-confocal) and 26% (confocal) in the Oka organ cells. This improved lentivirus expression system will provide us a useful tool for efficient gene transfer and expression in shrimp cells.

Single-run electrochemical determination of melamine in dairy products and pet foods.

A simple electrochemical approach, which does not require any expensive and complex instruments, is established for the selective and quantitative recognition of melamine in diary products and pet foods. During a preconcentration step (at 1.8 V versus Ag/AgCl), the formation of a polymer film from melamine on a preanodized screen-printed carbon electrode was identified by SEM and XPS. The as-formed polymer was found to be electroactive with a reversible redox peak, and hence square-wave voltammetry was applied to further increase the detection sensitivity to meet the detection limit for application in real sample analysis. Simply with a medium exchange procedure, melamine was selectively detected with a detection limit (S/N=3) of 0.8 µM (i.e., 98.3 ppb) by square-wave voltammetry. Lower than 1 ppm of melamine in real samples can be easily detected with good recoveries of 98.7-100.9% by the proposed approach. The recovery tests established for external calibration and standard addition techniques verified that the analysis can be done in a single-run measurement.