The detection and analysis of circulating tumor cells (CTCs) may enable a broad range of cancer-related applications, including the identification of acquired drug resistance during treatments. ...However, the non-scalable fabrication, prolonged sample processing times, and the lack of automation, associated with most of the technologies developed to isolate these rare cells, have impeded their transition into the clinical practice. This work describes a novel membrane-based microfiltration device comprised of a fully automated sample processing unit and a machine-vision-enabled imaging system that allows the efficient isolation and rapid analysis of CTCs from blood. The device performance was characterized using four prostate cancer cell lines, including PC-3, VCaP, DU-145, and LNCaP, obtaining high assay reproducibility and capture efficiencies greater than 93% after processing 7.5 mL blood samples spiked with 100 cancer cells. Cancer cells remained viable after filtration due to the minimal shear stress exerted over cells during the procedure, while the identification of cancer cells by immunostaining was not affected by the number of non-specific events captured on the membrane. We were also able to identify the androgen receptor (AR) point mutation T878A from 7.5 mL blood samples spiked with 50 LNCaP cells using RT-PCR and Sanger sequencing. Finally, CTCs were detected in 8 out of 8 samples from patients diagnosed with metastatic prostate cancer (mean ± SEM = 21 ± 2.957 CTCs/mL, median = 21 CTCs/mL), demonstrating the potential clinical utility of this device.
The COVID-19 pandemic has been the most critical public health issue in modern history due to its highly infectious and deathly potential, and the limited access to massive, low-cost, and reliable ...testing has significantly worsened the crisis. The recovery and the vaccination of millions of people against COVID-19 have made serological tests highly relevant to identify the presence and levels of SARS-CoV-2 antibodies. Due to its advantages, microfluidic-based technologies represent an attractive alternative to the conventional testing methodologies used for these purposes. In this work, we described the development of an automated ELISA on-chip capable of detecting anti-SARS-CoV-2 antibodies in serum samples from COVID-19 patients and vaccinated individuals. The colorimetric reactions were analyzed with a microplate reader. No statistically significant differences were observed when comparing the results of our automated ELISA on-chip against the ones obtained from a traditional ELISA on a microplate. Moreover, we demonstrated that it is possible to carry out the analysis of the colorimetric reaction by performing basic image analysis of photos taken with a smartphone, which constitutes a useful alternative when lacking specialized equipment or a laboratory setting. Our automated ELISA on-chip has the potential to be used in a clinical setting and mitigates some of the burden caused by testing deficiencies.
(OFI) contains health-promoting compounds like flavonoids, being the isorhamnetin glycosides the most abundant. We evaluated the effect of OFI extracts with different isorhamnetin glycosides against ...two different human colon cancer cells (HT-29 and Caco2). The extracts were obtained by alkaline hydrolysis with NaOH at 40 °C during 15, 30 or 60 min. Tri and diglycosides were the most abundant isorhamnetin glycosides, therefore these compounds were isolated to compare their cytotoxic effect with the obtained from the extracts. The OFI extracts and purified isorhamnetin glycosides were more cytotoxic against HT-29 cells than Caco2 cells. OFI-30 exhibited the lowest IC₅₀value against HT-29 (4.9 ± 0.5 μg/mL) and against Caco2 (8.2 ± 0.3 μg/mL). Isorhamnetin diglycosides IG5 and IG6 were more cytotoxic than pure isorhamnetin aglycone or triglycosides when they were tested in HT-29 cells. Bioluminescent analysis revealed increased activity of caspase 3/7 in OFI extracts-treated cells, particularly for the extract with the highest concentration of isorhamnetin triglycosides. Flow cytometry analysis confirmed that OFI extract and isorhamnetin glycosides induced a higher percentage of apoptosis in HT-29 than in Caco2, while isorhamnetin was more apoptotic in Caco2. This research demonstrated that glycosilation affected antiproliferative effect of pure isorhamnetin glycosides or when they are mixed with other phytochemicals in an extract obtained from OFI.
The presence of the genetic variants of the steroid 5-alpha reductase 2 enzyme, which is encoded by the
gene, has been associated with an increased risk of developing prostate cancer among certain ...ethnic groups. However, these molecular studies have not been conducted on the Mexican population. The analysis of the genetic variants, rs9282858 and rs523349, was performed in 101 males with prostate cancer and 100 healthy controls classified as males without prostate abnormalities (n=60) and males with benign prostatic hyperplasia (n=40), to identify a probable association with this cancer type in the Northeast Mexican population. An association was identified between prostate cancer and biomass exposure P=0.012; odds ratio (OR), 2.89; confidence interval (CI)=1.21-6.88 and tobacco use (P=0.028; OR=1.88; CI=1.07-3.31), while no association was observed between cancer development and the rs9282858 variant, or between a protective effect and the rs523349 variant. Notably, an association was identified between rs523349 and biomass exposure (P=0.013, OR=3.17; CI=1.23-8.17 for the G risk allele, and OR=0.32, CI=0.12-0.81 for the C protective allele) using the dominant genetic model. To the best of our knowledge, the present study was the first of its type to investigate the Mexican population with prostate cancer.
Circulating tumor cells (CTCs) have the potential of becoming the gold standard marker for cancer diagnosis, prognosis and monitoring. However, current methods for its isolation and characterization ...suffer from equipment variability and human operator error that hinder its widespread use. Here we report the design and construction of a fully automated high-throughput fluorescence microscope that enables the imaging and classification of cancer cells that were labeled by immunostaining procedures. An excellent agreement between our machine vision-based approach and a state-of-the-art microscopy equipment was achieved. Our integral approach provides a path for operator-free and robust analysis of cancer cells as a standard clinical practice.
We used continuous flow micro-devices as bioreactors for the production of a glycosylated pharmaceutical product (a monoclonal antibody). We cultured CHO cells on the surface of PMMA/PDMS ...micro-channels that had been textured by micromachining and coated with fibronectin. Three different micro-channel geometries (a wavy channel, a zigzag channel, and a series of donut-shape reservoirs) were tested in a continuous flow regime in the range of 3 to 6 μL min(-1). Both the geometry of the micro-device and the flow rate had a significant effect on cell adhesion, cell proliferation, and monoclonal antibody production. The most efficient configuration was a series of donut-shaped reservoirs, which yielded mAb concentrations of 7.2 mg L(-1) at residence times lower than one minute and steady-state productivities above 9 mg mL(-1) min(-1). These rates are at about 3 orders of magnitude higher than those observed in suspended-cell stirred tank fed-batch bioreactors.
Foams are high porosity and low density materials. In nature, they are a common architecture. Some of their relevant technological applications include heat and sound insulation, lightweight ...materials, and tissue engineering scaffolds. Foams derived from natural polymers are particularly attractive for tissue culture due to their biodegradability and bio-compatibility. Here, the foaming potential of an extensive list of materials was assayed, including slabs elaborated from whole flour, the starch component only, or the protein fraction only of maize seeds.
We used supercritical CO2 to produce foams from thermoplasticized maize derived materials. Polyethylene-glycol, sorbitol/glycerol, or urea/formamide were used as plasticizers. We report expansion ratios, porosities, average pore sizes, pore morphologies, and pore size distributions for these materials. High porosity foams were obtained from zein thermoplasticized with polyethylene glycol, and from starch thermoplasticized with urea/formamide. Zein foams had a higher porosity than starch foams (88% and 85%, respectively) and a narrower and more evenly distributed pore size. Starch foams exhibited a wider span of pore sizes and a larger average pore size than zein (208.84 vs. 55.43 μm2, respectively). Proof-of-concept cell culture experiments confirmed that mouse fibroblasts (NIH 3T3) and two different prostate cancer cell lines (22RV1, DU145) attached to and proliferated on zein foams.
We conducted screening and proof-of-concept experiments on the fabrication of foams from cereal-based bioplastics. We propose that a key indicator of foamability is the strain at break of the materials to be foamed (as calculated from stress vs. strain rate curves). Zein foams exhibit attractive properties (average pore size, pore size distribution, and porosity) for cell culture applications; we were able to establish and sustain mammalian cell cultures on zein foams for extended time periods.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Background Foams are high porosity and low density materials. In nature, they are a common architecture. Some of their relevant technological applications include heat and sound insulation, ...lightweight materials, and tissue engineering scaffolds. Foams derived from natural polymers are particularly attractive for tissue culture due to their biodegradability and bio-compatibility. Here, the foaming potential of an extensive list of materials was assayed, including slabs elaborated from whole flour, the starch component only, or the protein fraction only of maize seeds. Methodology/Principal Findings We used supercritical CO.sub.2 to produce foams from thermoplasticized maize derived materials. Polyethylene-glycol, sorbitol/glycerol, or urea/formamide were used as plasticizers. We report expansion ratios, porosities, average pore sizes, pore morphologies, and pore size distributions for these materials. High porosity foams were obtained from zein thermoplasticized with polyethylene glycol, and from starch thermoplasticized with urea/formamide. Zein foams had a higher porosity than starch foams (88% and 85%, respectively) and a narrower and more evenly distributed pore size. Starch foams exhibited a wider span of pore sizes and a larger average pore size than zein (208.84 vs. 55.43 mum.sup.2, respectively). Proof-of-concept cell culture experiments confirmed that mouse fibroblasts (NIH 3T3) and two different prostate cancer cell lines (22RV1, DU145) attached to and proliferated on zein foams. Conclusions/Significance We conducted screening and proof-of-concept experiments on the fabrication of foams from cereal-based bioplastics. We propose that a key indicator of foamability is the strain at break of the materials to be foamed (as calculated from stress vs. strain rate curves). Zein foams exhibit attractive properties (average pore size, pore size distribution, and porosity) for cell culture applications; we were able to establish and sustain mammalian cell cultures on zein foams for extended time periods.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
We report a proof-of-principle for the use of micro-devices as continuous bioreactors for the production of monoclonal antibodies. We culture CHO cells on the surface of PMMA "zigzag" channels ...textured with semi-spherical cavities coated with fibronectin, observing steady-state productivities 100 times higher than those observed in full scale systems.
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