Ratiometric fluorescent pH-sensitive polymers for high-throughput monitoring of extracellular pH

Zhang, Liqiang; Su, Fengyu; Kong, Xiangxing; Lee, Fred; Day, Kevin; Gao, Weimin; Vecera, Mary E.; Sohr, Jeremy M.; Buizer, Sean; Tian, Yanqing; Meldrum, Deirdre R.

doi:10.1039/c6ra06468j

Cited by 36 publications

(20 citation statements)

References 67 publications

(100 reference statements)

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“…AIE emission was detected at 520 nm and PtTFPP emission was measured at 650 nm. The measurement was carried out at 37 • C. Previously, we have showed that similar PHPMA copolymers were cell impermeable for a few cell lines, like E. coli and HeLa cell lines [63]. For avoiding the exchange of oxygen in the cell culture medium with air, a thin layer of mineral oil was added on the top of the LB medium.…”

Section: Cell Respirationmentioning

confidence: 99%

Poly(ε-caprolactone)-containing graft copolymers for ratiometric extracellular oxygen sensing

Zou¹,

Pan²,

Jiang³

et al. 2017

Sensors and Actuators B: Chemical

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a b s t r a c tA graft copolymer composed of poly(ε-caprolactone) as a hydrophobic chain and poly(N-(2-hydroxypropyl)methacrylamide) as a hydrophilic segment with an internal reference probe for oxygen sensing was prepared. Highly efficient platinum(II)meso-tetra(pentafluorophenyl)porphine (PtTFPP) was used as a typical oxygen probe. Although PtTFPP was highly efficient, it is hydrophobic, limiting its direct application for oxygen sensing in biological conditions. By using the amphiphilic graft copolymer, PTTFPP was successfully incorporated into the micelles formed by this polymer to not only enable the application of PtTFPP for biosensing in medium but also retain its high efficiency. The quantum efficiency of PtTFPP in the micelle solution under nitrogen can reach as high as 0.20. The PtTFPP-containing micelles were tested for high throughput cell respiration and bacteria detection. The influence of an antibiotic, antimycin as a representative example, on cell respiration was also studied. For achieving high accurate sensing, an internal built-in reference probe with aggregation induced emission properties were also polymerized in the graft copolymer. This study demonstrated that the graft polymer approach is a good way to enable highly efficient and hydrophobic oxygen probes for biosensing.

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Section: Cell Respirationmentioning

confidence: 99%

Poly(ε-caprolactone)-containing graft copolymers for ratiometric extracellular oxygen sensing

Zou¹,

Pan²,

Jiang³

et al. 2017

Sensors and Actuators B: Chemical

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“…With high accuracy, any suspension culture of mixed populations could be maintained in log phase growth for days in order to study transient invaders into microbial communities 33 or even microbiome system dynamics 34 . The advent of small molecule fluorescent reporters for metabolic fitness 35 , pH 36,37 , and CO2 38 , in addition to the hundreds of fluorescent protein sensors available to the synthetic biology community at large 39,40 , also impresses the seemingly unlimited potential of being able to multiplex and quantify changes in growth, gene expression, and the environment in real-time.…”

Section: Discussionmentioning

confidence: 99%

Flexible open-source automation for robotic bioengineering

Chory

Gretton

DeBenedictis

et al. 2020

Preprint

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INTRODUCTIONLiquid handling robots have become a biotechnology staple 1,2 , allowing laborious or repetitive protocols to be executed in highthroughput. However, software narrowly designed to automate traditional hand-pipetting protocols often struggles to harness the full capabilities of robotic manipulation. Here we present Pyhamilton, an open-source Python package that eliminates these constraints, enabling experiments that could never be done by hand. We used Pyhamilton to double the speed of automated bacterial assays over current software and execute complex pipetting patterns to simulate population dynamics. Next, we incorporated feedback-control to maintain hundreds of remotely monitored bacterial cultures in log-phase growth without user intervention. Finally, we applied these capabilities to comprehensively optimize bioreactor protein production by maintaining and monitoring fluorescent protein expression of nearly 500 different continuous cultures to explore the carbon, nitrogen, and phosphorus fitness landscape. Our results demonstrate Pyhamilton's empowerment of existing hardware to new applications ranging from biomanufacturing to fundamental biology. MAIN TEXTAutomation has been widely implemented in biotechnology 3 to facilitate routine tasks involved in DNA sequencing 4 , chemical synthesis 5 , drug discovery 6 , and molecular biology 7 . In principle, flexibly programmable robots could enable diverse experiments beyond the capabilities of human researchers, across a range of disciples within the sciences. Existing robotic software easily automates protocols designed for hand pipettes, but struggles to enable more specialized or sophisticated methods. As such, truly custom robot manipulation remains out of reach for most laboratories 2 , even those with well-established automation infrastructures.Bioautomation lags behind the rapidly advancing field of manufacturing, where robots are expected to be task-flexible, responsive to new situations, and interactive with humans or remote management systems when ambiguous situations or errors arise 2 . A key limitation is the lack of a comprehensive, suitably abstract, and accessible software ecosystem 8-10 . Though bioinformatics is becoming increasingly opensourced 11,12 , bioautomation has been slow to adopt key practices such as modularity, version control, and asynchronous programming.To address these issues, we developed Pyhamilton, a Python package that not only facilitates high-throughput operations within the laboratory, but also allows liquid-handling robots to execute previously unimaginable and increasingly impressive methods. With this package, users can use process scheduling, run simulations for experimental planning, implement error handling for straightforward troubleshooting, and easily integrate robots with external laboratory equipment. Design of Pyhamilton SoftwarePyhamilton enables Hamilton STAR and STARlet liquid handling robots to be programmed using standard Python. This allows for robotic method development to benefit from standa...

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“…This UPR response is activated to lower the misfolded or unfolded protein content. When ER stress becomes intense, the UPR response shifts to autophagic and pro-apoptotic signaling capabilities ending in cell death (32,33). To ascertain fidelity to correct protein folding, cells regulate protein-folding capacity in the ER according to need.…”

Section: Discussionmentioning

confidence: 99%

“…This means that curcumin's induction of the cellular antioxidant system and/or efficient mitophagy associated with restoration of ∆Ψm may push almost 50% of cells to go through the cell cycle and escape cell death. It is quite interesting to hypothesize that the alteration of lysosomes and activation/delocalization of TFEB may lead to mitochondrial biogenesis (33). In parallel, this is also associated with an increased level of mitophagy and overall this results in a restored quality of mitochondrial functions (33).…”

Section: Successful Curcumin-induced Autophagy Detected By Impedancemmentioning

confidence: 99%

“…It is quite interesting to hypothesize that the alteration of lysosomes and activation/delocalization of TFEB may lead to mitochondrial biogenesis (33). In parallel, this is also associated with an increased level of mitophagy and overall this results in a restored quality of mitochondrial functions (33). On the other hand, the anti-inflammatory action of curcumin has two aspects: the first is mediated through its ability to inhibit cyclooxygenase-2 (COX-2), lipoxygenase (LOX), and inducible nitric oxide synthase (iNOS) (49)(59, 60), and the second is mediated through the ability of curcumin to activate directly antioxidant enzymes, i.e., catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GSH-PX) (33,50).…”

Section: Successful Curcumin-induced Autophagy Detected By Impedancemmentioning

confidence: 99%

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Highlighting curcumin-induced crosstalk between autophagy and apoptosis: A biochemical approach coupling impedancemetry, imaging, and flow cytometry

Oyanguren

Rainey

Moustapha

et al. 2019

Preprint

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These authors contributed equally.Curcumin, a major active component of turmeric (Curcuma longa, L.), is known to have various effects on both healthy and cancerous tissues. In vitro studies suggest that curcumin inhibits cancer cell growth by activating apoptosis, but the mechanism underlying the anticancer effects of curcumin is still unclear. Since there is a consensus about endoplasmic reticulum (ER) stress being involved in the cytotoxicity of many natural compounds, we investigated by Amnis ® Imaging flow cytometry the mechanistic aspects of curcumin's destabilization of the ER, but also the status of the lysosomal compartment involved in curcumin-associated apoptosis. Curcumin induces ER stress thereby causing an unfolded protein response (UPR) and calcium release which destabilize the mitochondrial compartment and induce apoptosis. These events are also associated with secondary lysosomal membrane permeabilization and activation of caspase-8, mediated by activation of cathepsins and calpains. We previously showed that sequence lead to the generation of truncated tBid and disruption of mitochondrial homeostasis. These two pathways of different intensities and momentum converge towards an amplification of cell death that still needs to be studied in more detail. It has been suggested that it may be possible to exploit autophagy for cancer therapy. There is a complex interplay involving early autophagy as soon as mitochondria produce superoxide anions and hydrogen peroxide. Treatments with 10 µM to 20 µM curcumin induce autophagosome formation, while only early events of cell death are detectable.In the present study, curcumin-induced autophagy failed to rescue all cells since most cells underwent type II cell death following initial autophagic processes. However, a small number of cells blocked in the cell cycle escaped and were rescued to give rise to a novel proliferation phase. Curcumin | cell death | autophagy | apoptosis | ROS | Calcium | Stress | Mitochondria | Endoplasmic reticulum | cancer |

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Ratiometric fluorescent pH-sensitive polymers for high-throughput monitoring of extracellular pH

Cited by 36 publications

References 67 publications

Poly(ε-caprolactone)-containing graft copolymers for ratiometric extracellular oxygen sensing

Poly(ε-caprolactone)-containing graft copolymers for ratiometric extracellular oxygen sensing

Flexible open-source automation for robotic bioengineering

Highlighting curcumin-induced crosstalk between autophagy and apoptosis: A biochemical approach coupling impedancemetry, imaging, and flow cytometry

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