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Potential of alfalfa as an alternative molt induction diet for laying hens: egg quality and consumer acceptability.

Dietary molt induction to initiate additional egg laying cycles in commercial laying hen flocks is a wide spread practice in the United States. Feed deprivation is the most commonly used method but this practice has generated several concerns which has lead to research for viable alternative approaches. From a management standpoint a single ingredient molting diet consisting of high fiber-low energy represents an easily adaptable diet for large laying hen production units. Alfalfa meal is readily available in most commercial locations and possesses many of the desirable properties of an ideal laying hen molt diet. In the current study hens at a commercial laying facility were molted by both alfalfa and feed deprivation. After the hens had reentered post-molt commercial egg production, eggs were examined for egg quality performance. Egg shell strength, albumen height, yolk height, weight, length, and yolk color were all tested using various mechanical techniques. The eggs were also sampled for testing by consumer sensory panels that assessed the desirability of the eggs' color and flavor/texture. Eggs laid by hens molted by alfalfa had a significantly lower (p<0.05) "a*" level of colorimetry. Eggs laid by hens molted with alfalfa also exhibited significantly higher (p<0.05) egg weights and length. In the consumer sensory test, there was no significant difference (p>0.05) in color or flavor/texture scores in eggs from either feed deprived or alfalfa molted hens. The consumer sensory and mechanical quality attributes indicates that alfalfa shows promise as an alternative molt induction diet by providing a single diet option for extending egg production into a second egg laying cycle.

Animal Feed↗

Highly sensitive amperometric immunosensors for microcystin detection in algae.

The presence of cyanobacterial toxins in water and algae pose a health hazard for animals and humans, due to their tumour-promoting activity and carcinogen effects. The use of simple, rapid and reliable tools for routine analysis is becoming a necessity. With this purpose, our group has developed two electrochemical immunosensors for the detection of microcystin-LR (MC-LR) based on the affinity between this cyanotoxin and the corresponding monoclonal and polyclonal antibodies. A competitive direct enzyme-linked immunosorbent assays (ELISAs) was designed and, after validation of the approach on microtiter wells, screen-printed graphite electrodes were used as supports. Colorimetry was used to optimise the experimental parameters and to compare the performance of monoclonal and polyclonal antibodies. Afterwards, electrochemical measurements were performed at -200 mV (versus Ag/AgCl) using 5-methyl-phenazinium methyl sulfate (MPMS) as mediator for horseradish peroxidase (HRP), the enzymatic label of the competitor. The IC(50) values were 0.10 and 1.73 microgL(-1) for MAb and PAb, respectively. Whereas Mab provided higher sensitivities, the reproducibility was better when using PAb. The developed amperometric immunosensors were applied to the analysis of cyanobacterial samples from the Tarn River (Midi-Pyrénées, France) and the presence of MC was confirmed by the colorimetric protein phosphatase inhibition (PPI) assay and high performance liquid chromatography (HPLC). The limits of detection attained from the calibration curves and the results obtained for the real samples demonstrate the potential use of the immunosensors as screening tools for routine use in the assessment of water quality and the control of toxins in algae.

Biosensing Techniques↗

Interactions between 1-benzoyl-4-p-chlorophenyl thiosemicarbazide and serum albumin: investigation by fluorescence spectroscopy.

The interactions between 1-benzoyl-4-p-chlorphenyl thiosemicarbazide (BCPT) and bovine serum albumin (BSA) or human serum albumin (HSA) have been studied by fluorescence spectroscopy. By the analysis of fluorescence spectrum and fluorescence intensity, it was showed that BCPT has a strong ability to quench the intrinsic fluorescence of both bovine serum albumin and human serum albumin through a static quenching procedure. The binding constants of BCPT with BSA or HSA were determined at different temperatures based on the fluorescence quenching results. The binding sites were obtained and the binding force were suggested to be mainly hydrophobic. The effect of common ions on the binding constants was also investigated. A new fluorescence spectroscopy assay of the proteins is presented. The linear range is 5.36-67.0 microg mL(-1) with recovery of 101.1% for BSA, and the linear range is 8.28-144.9 microg mL(-1) with recovery of 102.6% for HSA. Determination of the proteins in bovine serum or in human serum by this method gives results which are very close to those obtained by using Coomassie Brilliant Blue G-250 colorimetry. A practical method was proposed for the determination of BCPT in human serum samples.

Animals↗

Peptide molecular lock-engineered nanobodies enable an oriented dual-modal immunoassay for reliable detection of Cronobacter sakazakii.

Conventional nanobody ELISAs for trace Cronobacter sakazakii in powdered infant formula suffer from random orientation and low signal output. We developed an oriented dual-modal immunoassay that combines site-specific biotinylation via a C-terminal AviTag and a peptide molecular lock, enabling controlled surface orientation while preserving nanobody structural integrity. This strategy was further integrated with phage-displayed nanobodies for multivalent amplification and both fluorescent and colorimetric readouts. The assay exhibited a broad linear range of 103-106&#xa0;CFU/mL, with limits of detection (LODs) of 6.70&#xa0;&#xd7;&#xa0;102&#xa0;CFU/mL for fluorescence and 1.55&#xa0;&#xd7;&#xa0;103&#xa0;CFU/mL for colorimetry, showing improved sensitivity compared with the conventional passive adsorption-based Nb-ELISA evaluated in this study. XGBoost-based multimodal fusion improved quantitative accuracy, and SHAP analysis elucidated modality contributions. In spiked powdered infant formula samples, recoveries ranged from 92.1% to 118% with coefficients of variation below 5.98%, confirming acceptable matrix tolerance and analytical reliability.

Cronobacter sakazakii↗

Determination of phenol and o-cresol by GC/MS in a fatal poisoning case.

A fatality due to the ingestion of solution containing phenol and o-cresol is described. The pathological findings were typical of acute substantial poisoning. Blood, urine and stomach content were obtained during post mortem examinations. Phenol and o-cresol were identified using GC/MS. The extractions from autopsy materials were obtained as follows: by gel permeation with cyclohexane/dichloromethane from stomach content, by solid phase extraction (SPE) from urine and by deproteinization with acetonitrile from blood. The phenol and o-cresol concentrations in the samples were found, respectively, as follows: 115.0 and 5.0 microg/g in the stomach contents, 58.3 and 1.9 microg/ml in the blood, 3.3 and 20.5 microg/ml in the urine. Distributions of phenol in fatal poisonings have been reported, but, usually, colorimetry was used as the analytical method and it cannot exclude the interference of other phenolic compounds.

Adult↗

Role of Smac/DIABLO in hydrogen peroxide-induced apoptosis in C2C12 myogenic cells.

Smac/DIABLO was recently identified as a protein released from mitochondria in response to apoptotic stimuli which promotes apoptosis by antagonizing inhibitors of apoptosis proteins. Furthermore, Smac/DIABLO plays an important regulatory role in the sensitization of cancer cells to both immune-and drug-induced apoptosis. However, little is known about the role of Smac/DIABLO in hydrogen peroxide (H(2)O(2))-induced apoptosis of C2C12 myogenic cells. In this study, Hoechst 33258 staining was used to examine cell morphological changes and to quantitate apoptotic nuclei. DNA fragmentation was observed by agarose gel electrophoresis. Intracellular translocation of Smac/DIABLO from mitochondria to the cytoplasm was observed by Western blotting. Activities of caspase-3 and caspase-9 were assayed by colorimetry and Western blotting. Full-length Smac/DIABLO cDNA and antisense phosphorothioate oligonucleotides against Smac/DIABLO were transiently transfected into C2C12 myogenic cells and Smac/DIABLO protein levels were analyzed by Western blotting. The results showed that: (1) H(2)O(2) (0.5 mmol/L) resulted in a marked release of Smac/DIABLO from mitochondria to cytoplasm 1 h after treatment, activation of caspase-3 and caspase-9 4 h after treatment, and specific morphological changes of apoptosis 24 h after treatment; (2) overexpression of Smac/DIABLO in C2C12 cells significantly enhanced H(2)O(2)-induced apoptosis and the activation of caspase-3 and caspase-9 (P<0.05). (3) Antisense phosphorothioate oligonucleotides against Smac/DIABLO markedly inhibited de novo synthesis of Smac/DIABLO and this effect was accompanied by decreased apoptosis and activation of caspase-3 and caspase-9 induced by H(2)O(2) (P<0.05). These data demonstrate that H(2)O(2) could result in apoptosis of C2C12 myogenic cells, and that release of Smac/DIABLO from mitochondria to cytoplasm and the subsequent activation of caspase-9 and caspase-3 played important roles in H(2)O(2)-induced apoptosis in C2C12 myogenic cells.

Animals↗

Assessing the role of race in quantitative measures of skin pigmentation and clinical assessments of photosensitivity.

BACKGROUND: Given the increasing demographic diversity in the United States, clarifying relationships between race, color, ethnicity, and disease processes is critical. OBJECTIVES: We sought to examine the correlation between objective measures of skin pigmentation, racial identification, and physician-diagnosed and self-reported skin phototypes. METHODS: A total of 558 participants (76 nonwhite) were evaluated. A subset underwent spectrometric readings and digital photography of the upper aspect of the inner arm. Self-identified race was compared with 7 measures of pigmentation. RESULTS: Race correlates best with physician-diagnosed skin phototype (r = 0.55, P < .01), whereas self-reported skin phototype, spectrometry, and colorimetry correlate poorly with race (r = 0.28, < 0.40, and r > -0.31, respectively, P < .01). Associations between race and subjective measures strengthen among patients with darker skin. CONCLUSION: Objective measures of pigmentation fail to correlate well with race, whereas race correlates moderately with physician-diagnosed skin phototype. Including objective methods of analyzing skin color may reduce subjective influences of race in assessing photosensitivity and potential risk for skin cancer.

Adult↗

Fluorescence spectroscopy studies on 5-aminosalicylic acid and zinc 5-aminosalylicylate interaction with human serum albumin.

The interaction between 5-aminosalicylic acid (5-ASA) or zinc 5-aminosalylicylate (5-ASZ) and human serum albumin (HSA) was studied by fluorescence spectroscopic technique. The binding constants of 5-ASA or 5-ASZ with HSA were determined at different temperatures under the optimum conditions. The binding sites were obtained and the acting force were suggested to be mainly hydrophobic. The effect of common ions on the binding constants was also investigated. A fluorescence spectroscopy assay of the proteins is presented in the paper. The determination results of the proteins in human serum by this method are very close to those obtained using Coomassie Brilliant Blue G-250 colorimetry, with relative standard deviations of 0.8-2.9%. A practical method was proposed for the determination of 5-ASA or 5-ASZ in human serum samples.

Humans↗

Selective quantitative bioanalysis of proteins in biological fluids by on-line immunoaffinity chromatography-protein digestion-liquid chromatography-mass spectrometry.

A quantitative method for the determination of proteins in complex biological matrices has been developed based on the selectivity of antibodies for sample purification followed by proteolytic digestion and quantitative mass spectrometry. An immunosorbent of polyclonal anti-bovine serum albumin (BSA) antibodies immobilized on CNBR agarose is used in the on-line mode for selective sample pretreatment. Next, the purified sample is trypsin digested to obtain protein specific peptide markers. Subsequent analysis of the peptide mixture using a desalination procedure and a separation step coupled, on-line to an ion-trap mass spectrometer, reveals that this method enables selective determination of proteins in biological matrices like diluted human plasma. This approach enhances substantially the selectivity compared to common quantitative analysis executed with immunoassays and colorimetry, fluorimetry or luminescence detection. Hyphenation of the immunoaffinity chromatography with on-line digestion and chromatography-mass spectrometry is performed and a completely on-line quantification of the model protein BSA in bovine and human urine was established. A detection limit of 170 nmol/l and a quantification limit of 280 nmol/l is obtained using 50 microl of either standard or spiked biological matrix. The model system allows fully automated absolute quantitative mass spectrometric analysis of intact proteins in biological matrices without time-consuming labeling procedures.

Animals↗

Determination of DPPH free radical scavenging activity by reversed-phase HPLC: a sensitive screening method for polyherbal formulations.

The colorimetric method of evaluation of antioxidant activity by scavenging 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical is with certain shortcomings like failure to indicate antioxidant activity of certain drugs and interference from color pigments of natural products. A specific HPLC method was developed for evaluating the DPPH free radical scavenging activity of commercial polyherbal formulations using a LiChrospher 100 RP-18e column (250 mm x 4 mm, 5 microM). The mobile phase was a mixture of methanol and water (80:20, v/v) pumped at a flow rate of 1 mL/min. The DPPH peaks were monitored at 517 nm. The method was standardized using known antioxidants such as ascorbic acid, 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (trolox), probucol and alpha-tocopherol. The 50% radical scavenging activity (IC50) determined by the HPLC method correlated well with that of colorimetry. This HPLC method was applied for the estimation of free radical scavenging activity of Silymarin and a few commercial hepatoprotective polyherbal formulations. While the colorimetric method failed to estimate the free radical scavenging activity of polyherbal formulations, HPLC method was free from interferences and was specific. The HPLC method is sensitive and can be used as a quality control tool for the rapid determination of free radical scavenging activity of variety of products including plant extracts, foods, drugs and polyherbal formulations.

Antioxidants↗

Antifibrotic effect of the Chinese herbs, Astragalus mongholicus and Angelica sinensis, in a rat model of chronic puromycin aminonucleoside nephrosis.

UNLABELLED: Nephrotic syndrome has long been treated in China with two herbs, Astragalus mongholicus and Angelica sinensis, which may have antifibrotic effects. METHODS: Rats with chronic puromycin-induced nephrosis were treated with Astragalus and Angelica 3 mL/d (n = 7) or enalapril 10 mg/kg/d (n = 7). Normal control rats (n = 7) received saline rather than puromycin, and an untreated control group (n = 7) received puromycin but no treatment. After 12 weeks, stained sections of the glomerulus and tubulointerstitium were evaluated for injury. Immunohistochemistry staining measured extracellular matrix components, transforming growth factor-beta1 (TGFbeta1), osteopontin, ED-1-positive cells, and alpha-actin. TGFbeta1 mRNA was assessed by in situ hybridization. Renin, ACE activity, angiotensin, and aldosterone were measured by radioimmunoassay or colorimetry. In the untreated rats, chronic renal injury progressed to marked fibrosis at 12 weeks. Astragalus and Angelica significantly reduced deterioration of renal function and histologic damage. Expressions of type III and IV collagen, fibronectin, and laminin also decreased significantly. This anti-fibrotic effect was similar to that of enalapril. The herbs had no effect on the renin-angiotensin system but did reduce the number of ED-1-positive, and alpha-actin positive cells and expression of osteopontin compared to untreated controls. The combination of Astragalus and Angelica retarded the progression of renal fibrosis and deterioration of renal function with comparable effects of enalapril. These effects were not caused by blocking the intrarenal renin-angiotensin system, but associated with suppression of the overexpression of TGFbeta1 and osteopontin, reduction of infiltrating macrophages, and less activation of renal intrinsic cells [corrected].

Angelica sinensis↗

Genetic risk for coronary artery disease in individuals with or without type 2 diabetes.

Given that a substantial proportion of individuals with coronary artery disease (CAD) also have type 2 diabetes, it is important to identify genes that confer susceptibility to CAD independently in subjects with type 2 diabetes and in those without this condition. A large-scale association study was performed to identify genes that confer susceptibility to CAD in either the absence or presence of type 2 diabetes. The study population comprised 5207 unrelated Japanese individuals, including 3085 subjects with CAD and 2122 controls. Among all subjects, 1704 individuals had type 2 diabetes and 3503 individuals did not have this condition. The genotypes for 33 polymorphisms of 27 candidate genes were determined with a fluorescence- or colorimetry-based allele-specific DNA primer-probe assay system. Multivariate logistic regression analysis with adjustment for age, body mass index, and the prevalence of smoking, hypertension, hypercholesterolemia, and hyperuricemia revealed that the following polymorphisms were significantly (P < 0.005) associated with CAD: the 1019C -->T of the connexin 37 gene for men with type 2 diabetes; the 2445G -->A in the fatty acid-binding protein 2 gene for women with this condition; the -863C-->A in the tumor necrosis factor-alpha gene, the -219G-->T in the apolipoprotein E gene, the 1019C-->T in the connexin 37 gene for men without type 2 diabetes; and the -482C-->T in the apolipoprotein C-III gene for women without this condition. Genotyping of these polymorphisms may prove informative for assessment of the genetic risk for CAD in the absence or presence of type 2 diabetes.

Aged↗

Culture of rat keratinocytes with acellular pig dermis.

This study evaluated the growth rate and the cell activity of cultured keratinocytes on acellular pig dermis in order to develop a composite skin in vitro for burn injuries or other skin defects. Full thickness skin was cultivated from neonatal SD rats, and separated into epidermal layer and dermal layer with enzyme digestion. The keratinocytes were then seeded on the prepared acellular pig dermis soaked in the culture medium. The cultures were incubated and the growth status of keratinocytes on acellular pig dermis evaluated by phase contrast microscope, histological examination with hematoxylin-eosin staining and acridine orange staining, immunohistochemistry, observation of growth curve plotted by MTT colorimetry and analysis of changes in keratinocytes proliferation cycle with flow cytometer. Almost all keratinocytes anchored in 48-72 h, and most inosculated at days 6 and 7. The growth curve showed that the keratinocytes grew in logarithmic phase at days 3-6 after seeding. More than four layers of keratinocyte structure and the basement membrane between keratinocytes and porcine dermis were observed. Pancytokeratin was strongly positive in the cultured keratinocytes. Laminin and collagen IV were positive in the basement membrane. It is concluded that the cultured keratinocytes on acellular pig dermis grow well and the structure of composite skin which has been established is satisfactory.

Animals↗

Reference ranges of electrolyte and anion gap on the Beckman E4A, Beckman Synchron CX5, Nova CRT, and Nova Stat Profile Ultra.

The widespread use of ion-selective electrode causes the reference range of the anion gap (AG) to be lowered from 8-16 to 3-11 mmol/l. The use of the outdated reference range (8-16 mmol/l) leads to the misinterpretation of the value of the anion gap. To interpret the anion gap accurately, one must use an analyzer-specific reference range. This study established the reference ranges of the electrolyte and anion gap in four ion-selective electrode analyzers. We collected clotted and lithium-heparinized blood from 124 healthy volunteers. We determined the electrolyte in the Beckman E4A (serum), Beckman Synchron CX5 (serum), and Nova CRT (serum and plasma). The anion gap was calculated from the formula: [Na(+)-(Cl(-)+HCO3(-))]. Blood sodium, potassium and bicarbonate were determined using the Nova Stat Profile Ultra. We used the plasma chloride from the Nova CRT to calculate the value of the anion gap in the Nova Stat Profile Ultra. We established the reference ranges using the non-parametric percentile estimation method. Accuracy and precision of the electrolyte performances obtained from all analyzers were acceptable. Reference values of serum and plasma sodium, potassium, and chloride were similar in all analyzers. The value of blood sodium obtained from the Nova Stat Profile Ultra was slightly higher than the values for the serum and plasma sodium obtained from the other analyzers. The bicarbonate ranges obtained from the Nova analyzers were higher than the values obtained from the Beckman analyzers. For the anion gap, the reference ranges in this study were low but similar to other studies (3-11 mmol/l) using ion-selective electrode. However, our reference ranges were lower than the previous reference ranges obtained from the continuous-flow analyzer (8-16 or 9-18 mmol/l) incorporated with flame photometry and colorimetry techniques.

Anions↗

The effect of hyperlipidemia on Technicon SMAC measurements.

The chemical measurements on our Technicon SMAC of lipemic sera before and after clearing lipemia by ultracentrifugation showed that uric acid, creatinine, carbon dioxide, calcium, phosphorus, potassium, and alkaline phosphatase were not affected significantly by lipemia, whereas sodium, urea, glucose, chloride and total protein showed small but significant increases with averages of less than 1.9 percent. Albumin showed a significant decrease of 1.2 percent. In contrast, the results for the enzymes, alanine aminotransferase (ALT), aspartate aminotransferase (AST) and lactate dehydrogenase (LDH) showed striking differences between pre- and post-centrifuged sera in a number of specimens. With lactate dehydrogenase, thirty-two of fifty specimens registered an increase in activity while with the aminotransferases, thirty-five and forty-one out of fifty specimens showed a decrease in aspartate aminotransferase and alanine aminotransferase activities, respectively. Although much of the lipemic interference can be explained by the volume displacement of serum by lipids or by interference by lipemia with colorimetry, the anomalous effects observed with the enzymes indicate the possibility of other, as yet, undetermined factor(s).

Autoanalysis↗

Oxidative stress may be a piece in the endometriosis puzzle.

OBJECTIVE: To assess the total antioxidant potential of women with endometriosis-associated infertility, women with idiopathic infertility, and fertile controls. DESIGN: Retrospective study. SETTING: University medical hospital. PATIENT(S): Sixty-five women admitted for diagnostic laparoscopy. MAIN OUTCOME MEASURES: Peritoneal fluid samples were analyzed for superoxide dismutase activity and total antioxidant status on spectrophotometry, glutathione peroxidase activity on enzyme-linked immunosorbent assay, and lipid peroxides levels on colorimetry. RESULT(S): Mean activity of superoxide dismutase, glutathione peroxidase, and total antioxidant status was lowest and lipid peroxide level was highest among infertile patients with endometriosis. Women with idiopathic infertility, in contrast, had the highest superoxide dismutase, glutathione peroxidase, and total antioxidant status activity and the lowest lipid peroxide level. CONCLUSION(S): High antioxidant potential is not a contributing factor in women with idiopathic infertility. Low total antioxidant status and low activity of antioxidant enzymes in the peritoneal fluid of infertile women with endometriosis probably do not influence fertility in these women, but these factors may play a role in the development of the disease.

Ascitic Fluid↗

Determination of inorganic cations and ammonium in environmental waters by ion chromatography with a high-capacity cation-exchange column.

While alkali and alkaline earth cations are commonly determined by using spectrometric techniques such as atomic absorption spectrometry or inductively coupled plasma, ammonium cation in the same sample must be measured separately by a wet chemical technique such as colorimetry, titrimetry, or ammonia-selective electrode. In a single 25-min run ion chromatography can determine all of the important inorganic cations including lithium, sodium, ammonium, potassium, magnesium and calcium. In this paper, we describe the use of ion chromatography with a new high-capacity cation-exchange column (the IonPac CS16), an electrolytically-generated methanesulfonic acid eluent and suppressed conductivity detection to determine dissolved alkali and alkaline earth cations and ammonium in drinking water wastewater and aqueous soil extracts. The IonPac CS16 is a high-capacity cation-exchange column that incorporates recent advances in polymer chemistry to enable trace-level determinations of cations even in high-ionic-strength matrices. We discuss the linear range, method detection limits, and analyte recoveries obtained with this column, and evaluate the effect of potential interferences on method performance during the analysis of typical environmental samples.

Cation Exchange Resins↗

Enzyme immunoassays for measuring complement-dependent prevention of immune precipitation (PIP) and solubilization of preformed antigen-antibody complexes (SOL).

We have developed simple and sensitive enzyme-based methods for evaluating the ability of serum complement to prevent immune complex precipitation (PIP) or to solubilize preformed immune complexes (SOL). Alkaline phosphatase, serving both as antigen and label, is added to goat IgG anti-alkaline phosphatase antibodies, with serum present throughout the assay (PIP), or added after immune complex formation (SOL). After incubation at 37 degrees C for 1 h followed by centrifugation, the enzyme activity of the supernatant, reflecting the amount of immune complexes in solution, is measured by colorimetry. Results are expressed with reference to a standard serum pool assigned 100 arbitrary units (AU). Intra- and inter-assay variabilities are within 10%. The normal ranges were 67-133 AU for PIP and 72-129 AU for SOL. These methods have been standardized for clinical use in relation to impaired complement function and immune complex disease, and adapted for measuring complement mediated binding of immune complexes to erythrocytes. They are sensitive, easy to perform and do not require expensive facilities. By measuring the interaction of complement with immune complexes, these methods may highlight aspects of the classical and the alternative pathway that are different from those detected using haemolysis as an endpoint.

Alkaline Phosphatase↗