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- Beal, Jacob, et al.
(författare)
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Robust estimation of bacterial cell count from optical density
- 2020
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Ingår i: Communications Biology. - : Springer Science and Business Media LLC. - 2399-3642. ; 3:1
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Tidskriftsartikel (refereegranskat)abstract
- Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data.
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4. |
- Zheng, Yiran
(författare)
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Exploring dual-lipopolysaccharide exposure on schizophrenia-like behavior and the kynurenine pathway in rodent models
- 2023
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Immune activation contributes to the pathophysiology of schizophrenia. The kynurenine pathway, serving as the primary route of tryptophan catabolism, has been posited as a potential link between immune activation and the manifestation of symptoms associated with schizophrenia. We previously demonstrated that the administration of dual-lipopolysaccharide (LPS) treatments induces the kynurenine pathway and elicits behavioral changes reminiscent of schizophrenia symptoms in mice. The dual-LPS treatment model may serve as a valuable tool for studying the effects of immune activation on the progression of schizophrenia. It is worth mentioning that rats and mice, which are frequently employed as laboratory animals in preclinical research, exhibit species-specific disparities in the kynurenine pathway metabolism. The present thesis aims to further characterize the behavioral effects induced by dual-LPS treatment in both mice and rats and evaluate the feasibility of the dual-LPS treatment model as a schizophrenia-like animal model. Moreover, we aim to investigate the changes in the kynurenine pathway in rats following dual-LPS treatment and explore possible species differences by comparing kynurenine pathway metabolism under physiological conditions and after dual-LPS treatment in both rats and mice. Thus, mice and rats administered dual-LPS treatment were studied for behavioral abnormalities associated with schizophrenia and biochemical changes in the kynurenine pathway metabolism in both the brain and periphery. Dual-LPS treatment induces behavioral changes associated with symptoms resembling schizophrenia in both rats and mice. In both species, dual-LPS treatment resulted in reduced spontaneous locomotion, heightened locomotion responsiveness to D-amphetamine, and the elicitation of anxiety-like behavior. In mice, dual-LPS treatment induced deficits in associative learning, while in rats, it was shown to impair recognition memory. Under basal physiological conditions, mice and rats exhibited distinct kynurenine pathway metabolite profiles in the brain, plasma, and liver. Rats consistently displayed higher KYNA/3HK and QUIN/PIC ratios across all brain regions, indicating a predominance of the neuroprotective branch in rats compared to mice under normal physiological conditions. In both mice and rats, dual-LPS treatment induced the kynurenine pathway, leading to elevated brain KYNA levels. Following dual-LPS treatment, species-specific changes were observed in brain, plasma, and liver tissues, with the most significant difference being the robust induction of QUIN in the mouse brain but not in the rat brain following dual-LPS treatment. Additionally, brain region-specific responses were observed in both species. Our radiochemical detection data confirmed that dual-LPS treatment alters the local kynurenine pathway metabolism in the brain, augmenting the de novo production of brain KYNA and increasing the KYNA/3-HK ratio in the striatum. The findings presented in this thesis further validate the utility of the dual-LPS model as a promising animal model of schizophrenia. Additionally, the results presented in the thesis comprehensively elucidate species-specific differences in kynurenine pathway metabolism under both physiological conditions and following dual-LPS treatment. These insights bear significant relevance for the selection of appropriate animal models when examining the kynurenine pathway, aiding in animal model selection for related research.
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