Lipid emulsion enhances fish allergen parvalbumin's resistance to in vitro digestion and IgG/IgE binding capacity.

This study was performed to determine Parvalbumin (PV), a well-known fish allergenic protein, digestion kinetics and immunoreactivity of digestion products with Immunoglobulin G/Immunoglobulin E recognition to understand its allergic potential with or without lipid emulsion process. PV was subjected to simulated gastrointestinal digestion in emulsified condition. Digestion kinetics of the protein was analysed by electrophoresis, IgG/IgE binding ability by immunoblotting and indirect ELISA. Lipid emulsion significantly (p < 0.01) reduced the degree of PV hydrolysis by 52.10% for gastric digestion. Immune fragments of gastric digestion were detectable for 90-120 min longer in emulsified condition showing resistance. Consequently, lipid emulsion decreased the digestive ability of PV in stomach, increasing resistance to gastrointestinal digestion by pepsin proteases. It also altered IgG/IgE binding ability of digestion products, thereby indicating that PV with lipid emulsion was resistant to digestion and possessed increased IgE binding ability resulting in higher risk of allergy among sensitized individuals.

Protein MRI contrast agent with unprecedented metal selectivity and sensitivity for liver cancer imaging.

With available MRI techniques, primary and metastatic liver cancers that are associated with high mortality rates and poor treatment responses are only diagnosed at late stages, due to the lack of highly sensitive contrast agents without Gd(3+) toxicity. We have developed a protein contrast agent (ProCA32) that exhibits high stability for Gd(3+) and a 10(11)-fold greater selectivity for Gd(3+) over Zn(2+) compared with existing contrast agents. ProCA32, modified from parvalbumin, possesses high relaxivities (r1/r2: 66.8 mmol(-1)⋅s(-1)/89.2 mmol(-1)⋅s(-1) per particle). Using T1- and T2-weighted, as well as T2/T1 ratio imaging, we have achieved, for the first time (to our knowledge), robust MRI detection of early liver metastases as small as ∼0.24 mm in diameter, much smaller than the current detection limit of 10-20 mm. Furthermore, ProCA32 exhibits appropriate in vivo preference for liver sinusoidal spaces and pharmacokinetics for high-quality imaging. ProCA32 will be invaluable for noninvasive early detection of primary and metastatic liver cancers as well as for monitoring treatment and guiding therapeutic interventions, including drug delivery.

Electrically induced transport of macromolecules through oral buccal mucosa.

OBJECTIVE: To investigate the feasibility of iontophoretic delivery of large molecules across buccal mucosa, and to establish its potential for enhanced drug delivery. METHODS: Qualitative (6h) and quantitative (8 and 36 h) assessment of porcine buccal mucosa, using a diffusion cell in vitro model, was carried out by fluorescent microscopy and UV/Vis spectroscopy respectively, with four fluorescently-labeled model species (3 and 10 kDa dextrans, 12 kDa parvalbumin and 66 kDa bovine serum albumin, BSA). Passive and iontophoresis parameters were obtained. The experimental iontophoresis data were compared with theoretical predictions. RESULTS: The two dextrans and parvalbumin showed enhanced permeation through buccal mucosa after anodal iontophoresis (1-6h). Passive diffusion and cathodal iontophoresis resulted in minimal permeation. BSA could not be measured by either mode. Iontophoretic delivery profiles compared to passive delivery, had reduced time lags (30-50 versus ~270 min) and increased flux (~37 times faster). Time lag factor/enhancement ratio (TLF/ER) data confirmed that iontophoresis significantly enhanced permeation. The diffusion coefficients (D, passive) for dextrans were significantly higher than for parvalbumin, with the converse obtained for solubility (C0); permeability coefficients (P) were similar for all three species. Potential differences (V) for the two higher kDa species were significantly higher than for the lowest kDa species. Experimental and theoretical data were in reasonable agreement. SIGNIFICANCE: The experimental and theoretical data, confirming enhanced delivery of the model species via iontophoresis, gave a suitable basis for its potential application in the mouth, in a clinical setting and opens pathways to further research for delivering precious drugs topically and systemically.

A comparative study of avidin-biotin-peroxidase complexes for the immunohistochemical detection of antigens in neural tissue.

It has been suggested that the use of avidin-biotin immunohistochemical techniques for antigen detection in neural tissue produces nonspecific background staining. For this reason neural tissue was used to test the quality, sensitivity and specificity of four commercially available antibody detection kits which use avidin or streptavidin binding to biotin. Free-floating, thick-section immunohistochemistry on perfusion fixed rat central nervous system revealed variability among staining kits for all parameters analyzed under the same experimental conditions. The reagents from the Vector 'Elite' kit were the most sensitive and specific, and received the highest overall rating for quality. Most commercial products tested could be used at greater dilutions than those recommended by the manufacturers without compromising specific staining. No staining was evident when the primary and secondary antibodies were omitted. This suggests that nonspecific binding is unlikely to be due to endogenous ligands, charge or hydrophilic reactions between these tertiary complexes and the tissue sections.