Development of a Time Projection Chamber Readout with Hybrid Pixel Sensors for Beam Monitoring.

To monitor the position and profile of therapeutic carbon beams in real-time, in this paper, we proposed a system called HiBeam-T. The HiBeam-T is a time projection chamber (TPC) with forty Topmetal-II- CMOS pixel sensors as its readout. Each Topmetal-II- has 72 × 72 pixels with the size of 83 µm × 83 µm. The detector consists of the charge drift region and the charge collection area. The readout electronics comprise three Readout Control Modules and one Clock Synchronization Module. This Hibeam-T has a sensitive area of 20 × 20 cm and can acquire the center of the incident beams. The test with a continuous 80.55 MeV/u 12C6+ beam shows that the measurement resolution to the beam center could reach 6.45 µm for unsaturated beam projections.

Towards Lightweight Neural Networks for Garbage Object Detection.

In recent years, garbage classification has become a hot topic in China, and legislation on garbage classification has been proposed. Proper garbage classification and improving the recycling rate of garbage can protect the environment and save resources. In order to effectively achieve garbage classification, a lightweight garbage object detection model based on deep learning techniques was designed and developed in this study, which can locate and classify garbage objects in real-time using embedded devices. Focusing on the problems of low accuracy and poor real-time performances in garbage classification, we proposed a lightweight garbage object detection model, YOLOG (YOLO for garbage detection), which is based on accurate local receptive field dilation and can run on embedded devices at high speed and with high performance. YOLOG improves on YOLOv4 in three key ways, including the design of DCSPResNet with accurate local receptive field expansion based on dilated-deformable convolution, network structure simplification, and the use of new activation functions. We collected the domestic garbage image dataset, then trained and tested the model on it. Finally, in order to compare the performance difference between YOLOG and existing state-of-the-art algorithms, we conducted comparison experiments using a uniform data set training model. The experimental results showed that YOLOG achieved AP0.5 of 94.58% and computation of 6.05 Gflops, thus outperformed YOLOv3, YOLOv4, YOLOv4-Tiny, and YOLOv5s in terms of comprehensive performance indicators. The network proposed in this paper can detect domestic garbage accurately and rapidly, provide a foundation for future academic research and engineering applications.

Two new selariscinins from Selaginella tamariscina (Beauv.) Spring.

Two new selariscinins named selariscinin F (1) and selariscinin G (2), along with one known selariscinin D (3) were isolated from Selaginella tamariscina. The structures of 1-3 were elucidated on the basis of chemical and spectral analysis, including 1D, 2D NMR analyses and HRESIMS.

Screening and identification of hepatotoxic component in Evodia rutaecarpa based on spectrum-effect relationship and UPLC-Q-TOFMS.

Evodia rutaecarpa (E. rutaecarpa) has been used to treat aches, vomiting and dysentery in traditional Chinese medicine. However, as a mildly toxic herb its toxic components have not been elucidated. An attempt was made to illuminate the hepatotoxic constituents of E. rutaecarpa. The 50% ethanol extracts of E. rutaecarpa from 19 different sources were used to establish UPLC fingerprints and administered to mice at a dose of 35 g/kg (crude medicine weight/mouse weight) once daily for 14 days. Serum levels of alanine transaminase, aspartate aminotransferase and liver coefficient were used as indices of liver injury. Additionally, the characteristic peaks of 19 fingerprints were identified. Spectrum-effect relationships between fingerprints and hepatotoxic indicators were analyzed using bivariate correlation analysis (BCA). The UPLC fingerprints were established and a total of 28 main compounds were identified. Because of the inherent variations in chemical compositions, the liver injury levels were different among the E. rutaecarpa samples from 19 sites of production. BCA results indicated that compounds dihydrorutaecarpine, 6-acetoxy-5-epilimonin, goshuyuamide I, 1-methyl-2-[(Z)-5-undecenyl]-4(1H)-quinolone, 1-methyl-2-[(4Z,7Z)-4,7-tridecadienyl]-4(1H)-quinolone, evocarpine and 1-methyl-2-[(6Z,9Z)-6,9-pentadecadienyl]-4(1H)-quinolone were tentatively determined as the primary hepatotoxic components. The present study provides a valuable method for the discovery of hepatotoxic constituents by combination of fingerprints and hepatotoxicity index.

Multi-analysis strategy for metabolism of Andrographis paniculata in rat using liquid chromatography/quadrupole time-of-flight mass spectrometry.

Compared with chemical drugs, it is a huge challenge to identify active ingredients of multicomponent traditional Chinese medicine (TCM). For most TCMs, metabolism investigation of absorbed constituents is a feasible way to clarify the active material basis. Although Andrographis paniculata (AP) has been extensively researched by domestic and foreign scholars, its metabolism has seldom been fully addressed to date. In this paper, high-performance liquid chromatography/quadrupole time-of-flight mass spectrometry was applied to analysis and characterization of AP metabolism in rat urine and feces samples after oral administration of ethanol extract. The differences in metabolites and metabolic pathways between the two biological samples were further compared. The chemical structures of 20 components were tentatively identified from drug-treated biological samples, including six prototype components and 14 metabolites, which underwent such main metabolic pathways as hydrolyzation, hydrogenation, dehydroxylation, deoxygenation, methylation, glucuronidation, sulfonation and sulfation. Two co-existing components were found in urine and feces samples, suggesting that some ingredients' metabolic processes were not unique. This study provides a comprehensive report on the metabolism of AP in rats, which will be helpful for understanding its mechanism.

Dynamic X-ray imaging with screen-printed perovskite CMOS array.

High performance X-ray detector with ultra-high spatial and temporal resolution are crucial for biomedical imaging. This study reports a dynamic direct-conversion CMOS X-ray detector assembled with screen-printed CsPbBr3, whose mobility-lifetime product is 5.2 × 10-4 cm2 V-1 and X-ray sensitivity is 1.6 × 104 µC Gyair-1 cm-2. Samples larger than 5 cm[Formula: see text]10 cm can be rapidly imaged by scanning this detector at a speed of 300 frames per second along the vertical and horizontal directions. In comparison to traditional indirect-conversion CMOS X-ray detector, this perovskite CMOS detector offers high spatial resolution (5.0 lp mm-1) X-ray radiographic imaging capability at low radiation dose (260 nGy). Moreover, 3D tomographic images of a biological specimen are also successfully reconstructed. These results highlight the perovskite CMOS detector's potential in high-resolution, large-area, low-dose dynamic biomedical X-ray and CT imaging, as well as in non-destructive X-ray testing and security scanning.

CsPbBr3-DMSO merged perovskite micro-bricks for efficient X-ray detection.

Inorganic perovskite wafers with good stability and adjustable sizes are promising in X-ray detection but the high synthetic temperature is a hindrance. Herein, dimethyl sulfoxide (DMSO) is used to prepare the CsPbBr3 micro-bricks powder at room temperature. The CsPbBr3 powder has a cubic shape with few crystal defects, small charge trap density, and high crystallinity. A trace amount of DMSO attaches to the surface of the CsPbBr3 micro-bricks via Pb-O bonding, forming the CsPbBr3-DMSO adduct. During hot isostatic processing, the released DMSO vapor merges the CsPbBr3 micro-bricks, producing a compact and dense CsPbBr3 wafer with minimized grain boundaries and excellent charge transport properties. The CsPbBr3 wafer shows a large mobility-lifetime (µτ) product of 5.16 × 10-4 cm2·V-1, high sensitivity of 14,430 µC·Gyair-1·cm-2, low detection limit of 564 nGyair·s-1, as well as robust stability in X-ray detection. The results reveal a novel strategy with immense practical potential pertaining to high-contrast X-ray detection. Electronic Supplementary Material: Supplementary material (further details of the characterization, SEM images, AFM images, KPFM images, schematic illustration, XRD patterns, XPS spectra, FTIR spectra, UPS spectra, and stability tests) is available in the online version of this article at 10.1007/s12274-023-5487-3.

The anticancer flavonoid chrysin induces the unfolded protein response in hepatoma cells.

Chrysin is a natural and biologically active flavonoid with anticancer effects. However, little is known about the adaptive response of cancer cells to chrysin. Chrysin reportedly has proteasome inhibitor activity. Previous studies demonstrated that proteasome inhibitors might induce endoplasmic reticulum (ER) stress response. In this study, we aimed to determine the effects of chrysin on hepatoma cells and roles of the ER-resident protein GRP78 (glucose-regulated protein 78) in its action. Also, we investigated the effects of green tea polyphenol (-)-epigallocatechin-3-gallate (EGCG), a natural GRP78 inhibitor, on the sensitivity of hepatoma cells to chrysin. Here, we report that chrysin inhibits hepatoma cells growth and induces apoptosis in a dose-dependent manner. Chrysin induces GRP78 overexpression, X-box binding protein-1 splicing and eukaryotic initiation factor 2α phosphorylation, hallmarks of the unfolded protein response. GRP78 knockdown potentiates chrysin-induced caspase-7 cleavage in hepatoma cells and enhances chrysin-induced apoptosis. EGCG overcomes chrysin-induced GRP78 expression. Combination of EGCG potentiates chrysin-induced caspase-7 and poly (ADP-ribose) polymerase (PARP) cleavage. Finally, EGCG sensitizes hepatoma cells to chrysin through caspase-mediated apoptosis. These data suggest that chrysin triggers the unfolded protein response. Abrogation of GRP78 induction may improve the anticancer effects of chrysin. Combination of EGCG and chrysin represents a new regimen for cancer chemoprevention and therapeutics.

The reciprocal regulation of gamma-synuclein and IGF-I receptor expression creates a circuit that modulates IGF-I signaling.

Insulin-like growth factor (IGF) system plays important roles in carcinogenesis and maintenance of the malignant phenotype. Signaling through the IGF-I receptor (IGF-IR) has been shown to stimulate the growth and motility of a wide range of cancer cells. γ-synuclein (SNCG) is primarily expressed in peripheral neurons but also overexpressed in various cancer cells. Overexpression of SNCG correlates with tumor progression. In the present study we demonstrated a reciprocal regulation of IGF-I signaling and SNCG expression. IGF-I induced SNCG expression in various cancer cells. IGF-IR knockdown or IGF-IR inhibitor repressed SNCG expression. Both phosphatidylinositol 3-kinase and mitogen-activated protein kinase were involved in IGF-I induction of SNCG expression. Interestingly, SNCG knockdown led to proteasomal degradation of IGF-IR, thereby decreasing the steady-state levels of IGF-IR. Silencing of SNCG resulted in a decrease in ligand-induced phosphorylation of IGF-IR and its downstream signaling components, including insulin receptor substrate (IRS), Akt, and ERK1/2. Strikingly, SNCG physically interacted with IGF-IR and IRS-2. Silencing of IRS-2 impaired the interaction between SNCG and IGF-IR. Finally, SNCG knockdown suppressed IGF-I-induced cell proliferation and migration. These data reveal that SNCG and IGF-IR are mutually regulated by each other. SNCG blockade may suppress IGF-I-induced cell proliferation and migration. Conversely, IGF-IR inhibitors may be of utility in suppressing the aberrant expression of SNCG in cancer cells and thereby block its pro-tumor effects.

Large-scale plasmid preparation for transient gene expression.

Large-scale transient gene expression of recombinant protein in mammalian cells requires a great amount of plasmid. An economical method for large-scale plasmid preparation, based on fed-batch fermentation and an improved plasmid extraction process, has been established. Fed-batch growth of E. coli was carried out in 5 l bioreactor by controlling the glucose concentration below 1 g l(-1) after the feeding was started. Plasmid yields of 490 and 580 mg l(-1) were achieved with two strains of E. coli cells bearing pCEP4-EGFP and pID-EG respectively, representing 24.5- and 26-fold increases over those of the batch culture in shake-flask. To improve the procedure for large-scale preparation of plasmid DNA, addition of RNase to resuspension buffer and ultrafiltration of clarified lysate were adopted, and the quality of the resultant plasmid was comparable to that of commercial kit as disclosed in the small-scale transient transfection. This plasmid production process has great potential in the large scale transient gene expression which needs a large quantity of plasmid DNA.

Research on serum metabolomics of ovariectomized rats and intervention effect of Cuscuta chinensis on metabolic pattern.

As a traditional Chinese medicine of invigorating the kidney, Cuscuta chinensis (CC) can be applied in improving the deficiency of kidney qi in menopausal women and regulating the level of estrogen. Previously, it was found that the ethanol extract of CC had an estrogen-like effect. In this study, the metabolic profile and metabolic pathways of rats in sham, ovariectomized model and CC groups were analyzed using UPLC-TOFMS-based metabolomics and the pattern recognition technology. The serum endogenouse metabolites could be well differentiated in different group, indicating significant differences of metabolic profiles. CC had an reverse adjustment effect on 14 differential metabolites of ovariectomized rats, including sinapyl alcohol, deoxycholic acid, prostaglandin B2, prostaglandin I2, dihydrosphingosine, choline, pentadecanoic acid, arachidonic acid, 1-stearoyl-Sn-Glycerol-3-Phosphocholine, palmitoleic acid, palmitic acid, vaccenic acid, oleic acid and stearic acid. Furthermore, these differential metabolites were categorized into several major pathways, such as biosynthesis of unsaturated fatty acids, lycerophospholipid metabolism and arachidonic acid metabolism. Therefore, it could be concluded that the estrogen-like effect of CC was related to the lipid metabolism to some extent. The research results provide useful help for the in-depth research and development of CC.

(-)-Epigallocatechin gallate sensitizes breast cancer cells to paclitaxel in a murine model of breast carcinoma.

INTRODUCTION: Paclitaxel (Taxol) is a microtubule-targeted agent that is widely used for cancer treatment. However, resistance to paclitaxel is frequently encountered in the clinic. There is increasing interest in identifying compounds that may increase the sensitivity to conventional chemotherapeutic agents. In this study, we investigated whether green tea polyphenol (-)-epigallocatechin gallate (EGCG) could sensitize breast carcinoma to paclitaxel in vivo. METHODS: Breast cancer cells were treated with or without EGCG and paclitaxel followed by detection of cell survival and apoptosis. c-Jun NH2-terminal kinase (JNK) phosphorylation and glucose-regulated protein 78 (GRP78) expression were detected by Western blotting. For in vivo study, 4T1 breast cancer cells were inoculated into Balb/c mice to establish a transplantation model. The tumor-bearing mice were treated with or without EGCG (30 mg/kg, i.p.) and paclitaxel (10 mg/kg, i.p.). Tumor growth was monitored. Apoptosis in tumor tissues was detected. Cell lysates from tumors were subjected to Western blot analysis of GRP78 expression and JNK phosphorylation. RESULTS: EGCG synergistically sensitized breast cancer cells to paclitaxel in vitro and in vivo. EGCG in combination with paclitaxel significantly induced 4T1 cells apoptosis compared with each single treatment. When tumor-bearing mice were treated with paclitaxel in combination with EGCG, tumor growth was significantly inhibited, whereas the single-agent activity for paclitaxel or EGCG was poor. EGCG overcame paclitaxel-induced GRP78 expression and potentiated paclitaxel-induced JNK phosphorylation in 4T1 cells both in vitro and in vivo. CONCLUSIONS: EGCG may be used as a sensitizer to enhance the cytotoxicity of paclitaxel.

[Studies on influence factors of gnsenoside Rg1 and Rb1 absorption in intestines of rats].

OBJECTIVE: To investigate the absorption of gnsenoside Rg1 and Rb1 in Radix Gngseng at different intestine segments of rats and the influence of the drug solution concentration, pH, P-gp inhibitor. METHOD: The intestine cannulation was performed for in situ recirculation. Gnsenoside Rg1, Rb1 and phenol red concentration in the flux were separately measured by the reversed phase HPLC and UV. RESULTS: When the concentration was raised from 0.075-0.75 g L(-1) and 0.03-0.3 g L(-1), the uptake of ginsenoside Rg1 and Rb1 was separately linearly increased (r >0.999), and no changes of K(a) absorption fraction and t(1/2) are found. The pH of flux has no effect on drug absorption. Ginsenoside Rg1 can be absorbed in the whole intestine and no changes of K(a), absorption fraction and t(1/2) refound and all the parameters of ginsenoside Rb1 at jejunum are higher than that at ileum and duodenum (P <0. 5). Further more, P-gp inhibitor verapamil has obvious effect on the intestinal absorption of ginsenoside Rb1 (P <0.5) while it has no effect on ginsenoside Rg1. CONCLUSION: The absorption of ginsenoside Rg1 and Rb1 in intestine of rat are first-order kinetics, the absorption mechanism is infered the passive diffusion. Ginsenoside Rg1 has no specific absorption locus in intestine of rat and ginsenoside Rb1 has specific absorption locus of jejunum. Meanwhile, ginsenoside Rb1 is the P-gp substrate, and could increase its fraction of bioavailability by corporation with P-gp inhibitor.

Characterization of estrogenic active ingredients in Cuscuta chinensis Lam. based on spectral characteristics and high­performance liquid chromatography/quadrupole time­of­flight mass spectrometry.

High-performance liquid chromatography (HPLC) is an efficient method that is widely used to assess the quality of traditional Chinese medicine (TCM). It is well known that the quality of TCM has a direct effect on its efficacy; therefore, in order to thoroughly explain how TCM exerts its efficacy, it is necessary to characterize its active ingredients and assess their quality. The application of the spectrum­effect method is crucial for determining the pharmacological basis of materials. The aim of the present study was to examine the correlation between chemical spectra and estrogenic activity of Cuscuta chinensis Lam., in order to reveal active compounds with potential therapeutic effects. The spectra of Cuscuta chinensis Lam. were recorded using HPLC, and estrogenic activity was determined using a uterus growth test and MTT assay. Combination of the results of bivariate analysis, principal component analysis and Gray relational analysis identified 19 active compounds, as follows: Quercetin­3­O­(2'­O­α­rhamnosy­6'­O­malony)­â€‹ß­D­glucoside, ka-empferol­3­O­ß­D­aplosyl­(1→2)­â€‹[­α­â€‹L­rhamnosy­(1→6)]­ß-wD-glucoside, 6­O­(E)­P­coumaroyl)­ß­â€‹D­fructofuranosyl­(2→1)­α­D­glucopyranoside, kaempferol­7­rhamnosy, kaempferol­3­ß­D-glucuronide, apigenin, 4­caffeoyl­5­coumaroylquinic acid, kaempferol­3­arabofuranoside, quercetin­3­O-ß­D-apiofuranosyl-(1→2)-ß­D­galactoside, dicaffeoylquinic acid, hyperin, quercitin, isorhamnetin, chlorogenic acid, quercetin, quercltrin­2''­gallate, quercetin­3, 7­α­L­dirhamnoside and stigmasterol, as well as one unknown compound. The present study laid a foundation for in vivo metabolic studies regarding Cuscuta chinensis Lam. and for the development of its clinical application.

High-density transient gene expression in suspension-adapted 293 EBNA1 cells.

Large-scale transient gene expression (TGE) in mammalian cells is an attractive method to rapidly produce recombinant proteins for pre-clinical studies, with some processes reported to reach 100 L. However, the yield remains low, hardly over 20 mg protein/L, mainly because the current TGEs have been performed at low cell density (approximately 5 x 10(5) cells/mL). In this study, the strategy to improve TGE focuses on facilitating transfection at high cell density. A high-density perfusion culture of 293 EBNA1 cells was established in 2-L bioreactor using Freestyle 293 expression medium (Invitrogen, Singapore) to grow the cells for transfection. Transfection was then carried out at 1 x 10(7) cells/mL using polyethylenimine (PEI) as DNA carrier, at the optimized conditions of 6 microg DNA/10(7) cells and 1:3 DNA to PEI mass ratio. During the post-transfection phase, 80.8 mg/L of the model protein, EPO was obtained at day 5.5 post-transfection (130 mg total EPO production) using a fed-batch culture mode. In comparison, perfusion cultures using an enriched SFM II medium resulted in a longer post-transfection production phase (8 days), and 227 mg of EPO was produced in 10.7 L medium, showing that high-density TGE enables the production of several hundreds of milligrams of protein in a 2 L bioreactor. In addition, a protocol for economical plasmid preparation based on anion exchange was also established to satisfy TGE's demand in terms of quality and quantity. To the best of our knowledge, this is the first report of transient transfections at a high cell density of up to 1 x 10(7) cells/mL.

Enhancement of hepatitis B surface antigen production by adenosine 5'-monophosphate in culture of recombinant Chinese hamster ovary cells.

Adenosine 5'-monophosphate was found effective in enhancing hepatitis B surface antigen (HBsAg) production in the culture of recombinant Chinese hamster ovary (CHO) cells, but it markedly inhibited cell growth. A culture strategy with periodical medium changes was established to increase HBsAg production and avoid the AMP-induced inhibition of cell growth. The enhanced energy supply might be one of the reasons for the increase in HBsAg production in the presence of AMP.

HPLC/Q-TOF-MS-Based Identification of Absorbed Constituents and Their Metabolites in Rat Serum and Urine after Oral Administration of Cistanche deserticola Extract.

As a famous health food in China, Cistanche deserticola (C. deserticola) suggested an estrogenic activity according to our previous study. However, no one clarifies its active material basis to date. To find more potentially active constituents and elucidate metabolic pathways of metabolites, a method to simultaneously analyze multiple absorbed constituents and metabolites from C. deserticola in rat serum and urine was established using high-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (HPLC/Q-TOF-MS). Based on HPLC/Q-TOF-MS method, a total of 24 components, involving 9 prototype constituents and 15 metabolites in rat serum and urine samples, were tentatively identified based on retention time, ultraviolet spectrum, MS data, compound fragmentation laws, published literatures, and reference substances. Most of the compounds existed in the form of metabolites. The proposed metabolic pathways of main metabolites were discussed, including methylation, demethylation, hydrolysis, hydroxylation, acetoxylation, glucuronidation, dehydrogenation, sulfation, esterification, and so on. Phenylethanoid glycosides were extensively metabolized and mutually transformed in vivo. This investigation provided valuable information for further study of the active ingredients and action mechanism of C. deserticola.

Determination of tricarboxylic acid cycle acids and other related substances in cultured mammalian cells by gradient ion-exchange chromatography with suppressed conductivity detection.

An ion-exchange chromatography method was established for simultaneously analyzing the tricarboxylic acid (TCA) cycle acids and other related substances in cultured mammalian cells, including citrate, cis-aconitate, isocitrate, alpha-ketoglutarate, succinate, malate, fumarate, oxaloacetate, trans-aconitate, phosphate, lactate and pyruvate. A Dionex 600 ion chromatograph with an ion suppressor and a conductivity detector, and an IonPac AS11-HC analytical column were employed. An NaOH gradient elution containing 13.5% methanol contributed to sufficient separation of target substances. The stability of carboxylic acids was investigated and oxaloacetate was found to be extremely unstable especially at pH 3. TCA cycle acids and other related substances in Chinese hamster ovary (CHO) cells were separated completely, and lactate, malate, phosphate, citrate and cis-aconitate were quantified due to their higher concentrations. In the quantification of the five substances, detection limits (S/N=3) ranged from 0.12 to 0.48 microM, the correlation coefficients from 0.9982 to 1.0000 in their linear ranges of concentration, and the recoveries from 87 to 95%. The metabolic status of CHO cells was analyzed on the basis of the intracellular concentrations of TCA cycle acids.

[Transient gene expression of soluble VEGFR2: I-IV].

The extracelluar domain I-IV of target gene VEGFR2 (Vascular endothelial growth factor receptor 2) was cloned from villus of trimester abortion by RT-PCR, and linked to the expression vectors. Then, the transfection conditions were optimized in serum-free suspension culture HEK293 using GFP (Green fluorescence protein) as the report gene. The results showed that the optimal transfection efficiency and cell number were obtained when the ratio of foreign DNA: PEI = 1:2 (W/W), DNA = 1.5 g /10(6) cells and shaking speed (120 r/min) in serum free medium in the beginning 4 hours of transfection. After optimizing the transfection conditions, the expression vector was successfully constructed for transient gene expression in HEK293, COS-7, and CHO-K1. The result shows that the target protein was only detected in CHO-K1 supernatant. Because of the C-terminal 8-His tag of target protein, target protein was subsequently purified using Ni2+-IDA and 5 mg purified protein was obtained in 1.5 L supernatant of CHOK1.

A cell-detaching reactor for inoculation of anchorage-dependent CHO and Vero cells between stepwise-expanded bioreactors.

A cell-detaching reactor was developed to collect cells growing on microcarriers for inoculation between stepwise-expanded bioreactors. It consisted of a trypsinization zone and a separation zone, which were separated by a 200-mesh stainless steel screen. The screen allowed the cells only to pass through to the next bioreactor, after the cells have been trypsinized and detached from microcarriers. The operating feasibility of the cell-detaching reactor was tested with anchorage-dependent recombinant Chinese hamster ovary (rCHO) and African green monkey kidney (Vero) cells. rCHO and Vero cells were first cultured in a small microcarrier bioreactor, and then inoculated via the cell-detaching reactor into either a packed-bed bioreactor (for rCHO cells) or a larger microcarrier bioreactor (for Vero cells). For rCHO cells, the cell density reached 1.3 x 10(7) cells/ml in the perfusion culture, and Vero cells reached 1.3 x 10(6) cells/ml in the batch culture.