The Bradford Method for Protein Quantitation

Kruger, Nicholas J.

doi:10.1007/978-1-60327-259-9_4

Cited by 344 publications

(382 citation statements)

References 12 publications

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“…Unlike EGFP•hSP‐C WT , the aberrantly processed products (arrows) in addition to the primary translational product (arrowhead) of the EGFP•hSP‐C I73T protein were preferentially localized at the plasma membrane. Biotinylated proteins were normalized by loading equal concentration of each protein sample (Bradford method (30)). D) Band intensities of plasma membrane expressed EGFP‐tagged total SP‐C products were quantified and results from at least three separate experiments are shown.…”

Section: Resultsmentioning

confidence: 99%

A Nonaggregating Surfactant Protein C Mutant Is Misdirected to Early Endosomes and Disrupts Phospholipid Recycling

Beers

Hawkins

Maguire

et al. 2011

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Interstitial lung disease in both children and adults has been linked to mutations in the lung-specific Surfactant protein C gene (SFTPC). Among these, the missense mutation (isoleucine to threonine at codon 73 = hSP-CI73T) accounts for ~30% of all described SFTPC mutations. We reported previously that unlike the BRICHOS misfolding SFTPC mutants, expression of hSP-CI73T induces lung remodeling and alveolar lipoproteinosis without a substantial ER stress response or ER-mediated intrinsic apoptosis. We show here that, in contrast to its wild type counterpart that is directly routed to lysosomal-like organelles for processing, SP-CI73T is misdirected to the plasma membrane and subsequently internalized to the endocytic pathway via early endosomes, leading to the accumulation of abnormally processed proSP-C isoforms. Functionally, cells expressing hSP-CI73T demonstrated both impaired uptake and degradation of surfactant phospholipid, thus providing a molecular mechanism for the observed lipid accumulation in patients expressing hSP-CI73T through the disruption of normal phospholipid recycling. Our data provide evidence for a novel cellular mechanism for conformational protein associated diseases, and suggest a paradigm for mistargeted proteins involved in the disruption of the endosomal/lysosomal sorting machinery.

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Section: Resultsmentioning

confidence: 99%

A Nonaggregating Surfactant Protein C Mutant Is Misdirected to Early Endosomes and Disrupts Phospholipid Recycling

Beers

Hawkins

Maguire

et al. 2011

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“…In brief, mouse neuronal cells treated with single AAA for 24 h were lysed using lysis buffer (Sigma‐Aldrich) according to the manufacturer's instructions. Protein concentrations were determined using Bradford's method (Kruger ). The total protein samples were loaded on sodium dodecyl sulfate–polyacrylamide gel electrophoresis and then transferred to polyvinylidene fluoride membranes.…”

Section: Methodsmentioning

confidence: 99%

Increasing carbohydrate availability in the hindgut promotes hypothalamic neurotransmitter synthesis: aromatic amino acids linking the microbiota–brain axis

Gao

et al. 2019

Journal of Neurochemistry

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The gut microbiota is increasingly recognized to modulate brain function by recent studies demonstrating the central effects of various gut microbial manipulation strategies. Our previous study demonstrated that antibiotic-induced alterations of hindgut microbiota are associated with changes in aromatic amino acid (AAA) metabolism and hypothalamic neurochemistry, while the underlying mechanistic insight is limited. Given that the microbial AAA metabolism can be affected by luminal carbohydrate availability, here we hypothesize that increasing hindgut carbohydrate availability affects the expression of neurotransmitters in the porcine hypothalamus. A hindgut microbiota-targeted strategy was adopted by increasing hindgut carbohydrate availability in a cecal-cannulated piglet model. Mechanistic involvement of AAAs along the gut microbiotaÀbrain axis was further investigated in mice and neuronal cells. Increasing carbohydrate availability by cecal starch infusion led to a decrease in hindgut AAA metabolism, and an increase in systemic AAA availability, central AAA-derived neurotransmitters (5-HT, dopamine), and neurotrophin BDNF in piglets, indicating that hindgut microbiota affect hypothalamic neurochemistry in an AAA-dependent manner. Single AAA i.p. injection in mice revealed that an increase in circulating tryptophan and tyrosine elevated their concentrations in brain and finally promoted the expressions of 5-HT, dopamine, and BDNF in a time-dependent manner. Neuronal cells treated with single AAAs in vitro further demonstrated that tryptophan and tyrosine enhanced 5-HT and dopamine synthesis, respectively, and promoted BDNF expression partly through the 5-HT1A/DRD1-CREB pathway. Our study reveals that increasing hindgut carbohydrate availability promotes hypothalamic neurotransmitter synthesis and that AAAs act as potential mediators between hindgut microbiota and brain neurochemistry.

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“…(). The difference in total protein content prior to and after precipitation of the antibody component with 12% polyethylene glycol (PEG; Sigma Aldrich) was determined by the Bradford method (Kruger ).…”

Section: Methodsmentioning

confidence: 99%

Can heat-killed Gordonia bronchialis enhance growth and immunity in rainbow trout (Oncorhynchus mykiss )?

et al. 2016

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In this study, the efficacy of heat-killed Gordonia bronchialis on growth performance, immune system and gastrointestinal structure in rainbow trout (Oncorhynchus mykiss) was evaluated. Fish (mean weight 30 g) were fed basal diet (control), or treatment diets containing 2.48 9 10 8 (low dose) or 1.24 9 10 9 (high dose) cells kg À1 feed of heat-killed G. bronchialis on a pulse basis (5 days on treatment diet; 10 days on basal diet) for 95 days. On days 95 and 105, some of the fish were sampled for analysis of mentioned parameters. On days 110 and 120, the remaining fish were injected intraperitoneally with a 20 mL L À1 suspension of chicken red blood cells. Results showed that growth performance was significantly enhanced in both treatment groups compared with the control group. Serum complement and lysozyme activities and hemagglutination antibody titre were higher in both treatment groups compared with the control group. The length of the intestinal and pyloric caeca folds was increased in the high-dose group. Meanwhile, the number of goblet cells was increased in both treatment groups. This study suggests that heat-killed G. bronchialis has the potential to enhance growth, immunological parameters and the gastrointestinal structure in rainbow trout.

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The Bradford Method for Protein Quantitation

Cited by 344 publications

References 12 publications

A Nonaggregating Surfactant Protein C Mutant Is Misdirected to Early Endosomes and Disrupts Phospholipid Recycling

A Nonaggregating Surfactant Protein C Mutant Is Misdirected to Early Endosomes and Disrupts Phospholipid Recycling

Increasing carbohydrate availability in the hindgut promotes hypothalamic neurotransmitter synthesis: aromatic amino acids linking the microbiota–brain axis

Can heat-killed Gordonia bronchialis enhance growth and immunity in rainbow trout (Oncorhynchus mykiss )?

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