Tsutomu Honjoh

Correlation between plasma leptin concentration and body fat content in dogs

OBJECTIVE To evaluate the relationship between plasma leptin concentration and body fat content in dogs. ANIMALS 20 spayed female Beagles that were 10 months old at the start of the experiment. PROCEDURE Dogs were kept under regulated feeding and exercise conditions for 21 weeks, resulting in a wide range of body weights, body condition scores (BCS), and subcutaneous thicknesses. Plasma leptin concentration was measured by use of a canine leptin-specific ELISA test to evaluate its correlation to body fat content estimated by the deuterium oxide dilution method. Plasma concentrations of glucose, cholesterol, triglycerides (TG), and nonesterified fatty acids (NEFA) were also measured. RESULTS Body fat content (9 to 60% of body weight) was positively and closely correlated (r = 0.920; n = 20; P < 0.001) to plasma leptin concentration (0.67 to 8.06 ng/ml), compared with other variables (ie, glucose, cholesterol, TG, and NEFA; r = 0.142, 0.412, 0.074, and 0.182, respectively). CONCLUSIONS AND CLINICAL RELEVANCE The positive relationship between plasma leptin concentration and body fat content in dogs was similar to correlations reported for humans and rodents, suggesting that plasma leptin is a quantitative marker of adiposity in dogs.

A glass fiber sheet-based electroosmotic lateral flow immunoassay for point-of-care testing.

We have developed a quantitative immunoassay chip targeting point-of-care testing. To implement a lateral flow immunoassay, a glass fiber sheet was chosen as the material for the microfluidic channel in which the negative charge on the fiber surfaces efficiently generates the electroosmotic flow (EOF). The EOF, in turn, allows controllable bound/free separation of antigen/antibody interactions on the chip and enables precise determination of the antigen concentration. In addition, the defined size of the porous matrix was suitable for the filtration of undesired large particles. We confirmed the linear relationship between the concentration of analyte and the resulting fluorescence intensity from the immunoassay of two model analytes, C-reactive protein (CRP) and insulin, demonstrating that analyte concentration was quantitatively determined within the developed chip in 20 min. The limits of detection were 8.5 ng mL(-1) and 17 ng mL(-1) for CRP and insulin, respectively.

A harmonized immunoassay with liquid chromatography-mass spectrometry analysis in egg allergen determination

AbstractFood allergy is a serious health issue worldwide. Implementing allergen labeling regulations is extremely challenging for regulators, food manufacturers, and analytical kit manufacturers. Here we have developed an “amino acid sequence immunoassay” approach to ELISA. The new ELISA comprises of a monoclonal antibody generated via an analyte specific peptide antigen and sodium lauryl sulfate/sulfite solution. This combination enables the antibody to access the epitope site in unfolded analyte protein. The newly developed ELISA recovered 87.1%–106.4% ovalbumin from ovalbumin-incurred model processed foods, thereby demonstrating its applicability as practical egg allergen determination. Furthermore, the comparison of LC-MS/MS and the new ELISA, which targets the amino acid sequence conforming to the LC-MS/MS detection peptide, showed a good agreement. Consequently the harmonization of two methods was demonstrated. The complementary use of the new ELISA and LC-MS analysis can offer a wide range of practical benefits in terms of easiness, cost, accuracy, and efficiency in food allergen analysis. In addition, the new assay is attractive in respect to its easy antigen preparation and predetermined specificity. Graphical abstractThe ELISA composing of the monoclonal antibody targeting the amino acid sequence conformed to LC-MS detection peptide, and the protein conformation unfolding reagent was developed. In ovalbumin determination, the developed ELISA showed a good agreement with LC-MS analysis. Consequently the harmonization of immunoassay with LC-MS analysis by using common target amino acid sequence was demonstrated.