A novel fluorimetric assay for visualization and quantification of protein carbonyls in muscle foods.

A novel fluorimetric assay for visualization and quantification of protein carbonyls in muscle foods.
Muscle meals, notably fish merchandise are extremely uncovered to oxidative stress throughout processing and storage, leading to oxidative modification of proteins. Protein carbonyls content material has been used as one of many measures of oxidative stress.
Usually, the ensuing carbonylated proteins (CPs) have to date been labeled with 2,4-dinitrophenyl (DNP) hydrazine and detected with anti-DNP antibody. Nonetheless, the applicability of this technique to meals samples is proscribed by its excessive worth, time-consuming process and risk to carry out the measurements simply on soluble protein fractions.
We developed an easier, quicker and cheaper technique to evaluate CP degree in muscle meals, together with each soluble and insoluble protein fractions, which is predicated on a direct response of protein carbonyls with 7-(diethylamino)coumarin-3-carbohydrazide (CHH). The paper describes a novel approach to label each soluble and insoluble carbonylated proteins with CHH and decide carbonyl content material by fluorescence microscopy assay which correlates (R = 0.911) with typical ELISA technique.

Protein carbonyl willpower by a rhodamine B hydrazide-based fluorometric assay.

A brand new fluorometric assay is introduced for the ultrasensitive quantification of complete protein carbonyls, and is predicated on their particular response with rhodamine B hydrazide (RBH), and the manufacturing of a protein carbonyl-RBH hydrazone the fluorescence of which (at ex/em 560/585 nm) is enormously enhanced by guanidine-HCl.
In comparison with the fluorescein-5-thiosemicarbazide (FTC)-based fluorometric assay, the RBH assay makes use of a 24-fold shorter response incubation time (1 h) and at the very least 1000-fold decrease protein amount (2.5 µg), and produces very dependable information that had been verified by intensive standardization experiments.
The protein carbonyl group detection sensitivity restrict of the RBH assay, based mostly on its commonplace curve, will be as little as 0.Four pmol, and even decrease. Counting the very low protein restrict of the RBH assay, its cumulative and purposeful sensitivity is 8500- and 800-fold larger than the corresponding ones for the FTC assay. Neither heme proteins hemoglobin and cytochrome c nor DNA intervene with the RBH assay.

Protein and cell wall polysaccharide carbonyl willpower by a impartial pH 2,4-dinitrophenylhydrazine-based photometric assay.

A brand new 2,4-dinitrophenylhydrazine (DNPH)-based photometric assay is developed for the quantification of carbonyls in protein samples from any organic supply by protein carbonyl-DNPH hydrazone formation at acidic pH within the presence of denaturing urea, and subsequent hydrazone solubilization within the presence of SDS and stabilization from acid hydrolysis at pH 7.0.
At this impartial (ntr) pH, interfering unreacted DNPH is uncharged and its thus elevated hydrophobicity permits its 100% efficient removing from the solubilizate with ethyl acetate/hexane wash. The ntrDNPH assay is extra dependable and delicate than the usual (std) DNPH photometric assay as a result of it eliminates its essential limitations: (i) interfering unreacted DNPH (pKa 1.55) that’s nonspecifically certain to the TCA (pKa 0.7)-protein pellet is just not successfully eliminated after wash with EtOH: ethyl acetate as a result of it’s positively charged, (ii) acid (TCA-induced) hydrolysis of the protein carbonyl-DNPH hydrazone, (iii) pattern protein focus re-determination, (iv) lack of pattern acid (TCA)-soluble proteins, (v) DNA interference, and (vi) requires excessive protein amount samples (≥ 1 mg). Contemplating ntrDNPH assay’s very low protein restrict (1 µg), its cumulative and purposeful sensitivities are 2600- and 2000-fold larger than these of the stdDNPH assay, respectively.
The current research elucidates the DNA interference mechanism on the stdDNPH assay, and in addition develops a standardized protocol for pattern protein therapy and fractionation (into cytoplasmic/aqueous, membrane/lipid-bound, and histone/DNA-bound proteins; see Complement part V) to be able to guarantee reproducible carbonyl willpower on outlined cell protein fractions, and to remove assay interference from protein samples containing (i) Cys sulfenic acid teams (through their neutralization with dithiothreitol), and (ii) DNA (through its removing by streptomycin sulfate precipitation).
Lastly, the ntrDNPH assay determines carbonyl teams on cell wall polysaccharides, thus paving the best way on research to analyze cell partitions appearing as antioxidant protection in crops, fungi, micro organism and lichens.

Assay of urinary protein carbonyl content material can predict the steroid dependence and resistance in kids with idiopathic nephrotic syndrome.

Nephrotic syndrome in pediatric age is generally idiopathic. Idiopathic nephrotic syndrome (INS) by default is handled with steroids from the very starting. Some don’t reply to steroids and are grouped later as both steroid-resistant (SR) or steroid-dependent (SD) circumstances.
The protein selectivity index usually fails to foretell the SR and SD circumstances. The SD and SR circumstances of INS exhibit larger levels of oxidative stress in comparison with steroid responders. Proteins get carbonylated when they’re uncovered to free radicals. The importance of excretion of those carbonylated proteins in urine is but to be studied intimately. On this research, 70 circumstances of INS had been enrolled, and urinary protein carbonyl content material (UPCC) was estimated by Levine’s technique earlier than beginning the steroid remedy.
All of the circumstances had been adopted up and, based mostly on the response to steroid remedy, had been grouped as Group A (n = 47). Steroid delicate and Group B (n = 23), SD + SR circumstances. UPCC was considerably larger in Group B in comparison with Group A. Receiver-operating curve confirmed at a cutoff restrict of 5.10 nmoles/mg of protein, UPCC can predict SD or SR circumstances with 83.3% sensitivity and 85.2% specificity and space below the curve of 0.833, P<0.05. UPCC ranges greater than 5.10 nmoles/mg of protein, earlier than beginning the remedy can predict SD or SR in pediatric INS circumstances.

Fluorimetric screening assay for protein carbonyl analysis in organic samples.

Many assays can be found for the detection of protein carbonyls (PCs). At the moment, the measurement of PC teams after their derivatization with 2,4-dinitrophenol hydrazine (DNPH) is extensively used for measuring protein oxidation in organic samples.
Nonetheless, this technique contains a number of washing steps. On this context, we’ve developed a speedy, delicate, and correct fluorimetric technique tailored to 96-well microplates for the handy evaluation of protein carbonyl degree in organic samples.
The tactic reported right here is predicated on the response of carbonyl content material in proteins with 7-hydrazino-4-nitrobenzo-2,1,3-oxadiazole (NBDH) to kind extremely fluorescent derivatives through hydrazone formation. PCs had been decided utilizing the DNPH and NBDH assays in totally decreased bovine serum albumin (BSA) and plasma and liver homogenates obtained from wholesome management rats up the addition of varied quantities of HOCl-oxidized BSA (OxBSA).

Protein Carbonyl Content Assay Kit

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Description: Assay Kit for detection of Protein Carbonyl Content in the research laboratory

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CheKine™ Micro Protein Carbonyl Assay Kit

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Description: Abbkine CheKine™ Micro Protein Carbonyl Assay Kit is designed for detecting Protein Carbonyl content in the sample.

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OxiSelect Protein Carbonyl Fluorometric Assay, Trial Size

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Description: The most common products of protein oxidation in biological samples are the carbonyl derivatives of proline, lysine, arginine and threonine residues. Such derivatives are chemically stable and serve as markers for oxidative stress in most types of reactive oxygen species. Our OxiSelect Protein Carbonyl Fluorometric Assay Kit provides a rapid, efficient method for the detection of protein carbonyl residues. The fluorescence plate-based format provides a convenient system for direct measurement of protein carbonyl content.

Total Carbonyl (CO) Assay Kit

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Protein Carbonyl Colorimetric Assay Kit -Tissue and Serum Samples-

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Protein Carbonyl Colorimetric Assay Kit -Tissue and Serum Samples-

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Rat Protein Carbonyl ELISA kit

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Rat Protein Carbonyl ELISA kit

E02P0048-48 1 plate of 48 wells
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Description: A competitive ELISA for quantitative measurement of Rat Protein Carbonyl in samples from blood, plasma, serum, cell culture supernatant and other biological fluids. This is a high quality ELISA kit developped for optimal performance with samples from the particular species.

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Dog Protein Carbonyl ELISA kit

E08P0048-192T 192 tests
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Description: A competitive ELISA for quantitative measurement of Canine Protein Carbonyl in samples from blood, plasma, serum, cell culture supernatant and other biological fluids. This is a high quality ELISA kit developped for optimal performance with samples from the particular species.

Dog Protein Carbonyl ELISA kit

E08P0048-48 1 plate of 48 wells
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Description: A competitive ELISA for quantitative measurement of Canine Protein Carbonyl in samples from blood, plasma, serum, cell culture supernatant and other biological fluids. This is a high quality ELISA kit developped for optimal performance with samples from the particular species.

Dog Protein Carbonyl ELISA kit

E08P0048-96 1 plate of 96 wells
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Description: A competitive ELISA for quantitative measurement of Canine Protein Carbonyl in samples from blood, plasma, serum, cell culture supernatant and other biological fluids. This is a high quality ELISA kit developped for optimal performance with samples from the particular species.

Pig Protein Carbonyl ELISA kit

E07P0048-192T 192 tests
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Description: A competitive ELISA for quantitative measurement of Porcine Protein Carbonyl in samples from blood, plasma, serum, cell culture supernatant and other biological fluids. This is a high quality ELISA kit developped for optimal performance with samples from the particular species.

Pig Protein Carbonyl ELISA kit

E07P0048-48 1 plate of 48 wells
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Description: A competitive ELISA for quantitative measurement of Porcine Protein Carbonyl in samples from blood, plasma, serum, cell culture supernatant and other biological fluids. This is a high quality ELISA kit developped for optimal performance with samples from the particular species.

Pig Protein Carbonyl ELISA kit

E07P0048-96 1 plate of 96 wells
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Description: A competitive ELISA for quantitative measurement of Porcine Protein Carbonyl in samples from blood, plasma, serum, cell culture supernatant and other biological fluids. This is a high quality ELISA kit developped for optimal performance with samples from the particular species.

Goat Protein Carbonyl ELISA kit

E06P0048-192T 192 tests
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Description: A competitive ELISA for quantitative measurement of Goat Protein Carbonyl in samples from blood, plasma, serum, cell culture supernatant and other biological fluids. This is a high quality ELISA kit developped for optimal performance with samples from the particular species.

Goat Protein Carbonyl ELISA kit

E06P0048-48 1 plate of 48 wells
EUR 624
Description: A competitive ELISA for quantitative measurement of Goat Protein Carbonyl in samples from blood, plasma, serum, cell culture supernatant and other biological fluids. This is a high quality ELISA kit developped for optimal performance with samples from the particular species.

Goat Protein Carbonyl ELISA kit

E06P0048-96 1 plate of 96 wells
EUR 822
Description: A competitive ELISA for quantitative measurement of Goat Protein Carbonyl in samples from blood, plasma, serum, cell culture supernatant and other biological fluids. This is a high quality ELISA kit developped for optimal performance with samples from the particular species.

Mouse Protein Carbonyl ELISA kit

E03P0048-192T 192 tests
EUR 1524
Description: A competitive ELISA for quantitative measurement of Mouse Protein Carbonyl in samples from blood, plasma, serum, cell culture supernatant and other biological fluids. This is a high quality ELISA kit developped for optimal performance with samples from the particular species.

Mouse Protein Carbonyl ELISA kit

E03P0048-48 1 plate of 48 wells
EUR 624
Description: A competitive ELISA for quantitative measurement of Mouse Protein Carbonyl in samples from blood, plasma, serum, cell culture supernatant and other biological fluids. This is a high quality ELISA kit developped for optimal performance with samples from the particular species.

Mouse Protein Carbonyl ELISA kit

E03P0048-96 1 plate of 96 wells
EUR 822
Description: A competitive ELISA for quantitative measurement of Mouse Protein Carbonyl in samples from blood, plasma, serum, cell culture supernatant and other biological fluids. This is a high quality ELISA kit developped for optimal performance with samples from the particular species.

Human Protein Carbonyl ELISA kit

E01P0048-192T 192 tests
EUR 1524
Description: A competitive ELISA for quantitative measurement of Human Protein Carbonyl in samples from blood, plasma, serum, cell culture supernatant and other biological fluids. This is a high quality ELISA kit developped for optimal performance with samples from the particular species.

Human Protein Carbonyl ELISA kit

E01P0048-48 1 plate of 48 wells
EUR 624
Description: A competitive ELISA for quantitative measurement of Human Protein Carbonyl in samples from blood, plasma, serum, cell culture supernatant and other biological fluids. This is a high quality ELISA kit developped for optimal performance with samples from the particular species.

Human Protein Carbonyl ELISA kit

E01P0048-96 1 plate of 96 wells
EUR 822
Description: A competitive ELISA for quantitative measurement of Human Protein Carbonyl in samples from blood, plasma, serum, cell culture supernatant and other biological fluids. This is a high quality ELISA kit developped for optimal performance with samples from the particular species.
Utilizing the NBDH assay, PC concentrations as little as 0.2 nmol/mg had been detected with precision as little as 5%. Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectroscopy was used to efficiently establish the formation of the NBDH adducts after derivatization with commonplace oxidized peptides. Lastly, the 2 strategies had been additional used for PC willpower in plasma and liver samples from diabetic and regular rats, exhibiting that the NBDH assay will be reliably utilized in organic experiments.

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