Hapten Synthesis and Monoclonal Antibody Preparation for Simultaneous Detection of Albendazole and Its Metabolites in Animal-Origin Food

Albendazole (ABZ) is one of the benzimidazole anthelmintics, and the overuse of ABZ in breeding industry can lead to drug resistance and a variety of toxic effects in humans. Since the residue markers of ABZ are the sum of ABZ and three metabolites (collectively referred to as ABZs), albendazole-sulfone (ABZSO2), albendazole-sulfoxide (ABZSO), and albendazole-2-amino-sulfone (ABZNH2SO2), an antibody able to simultaneously recognize ABZs with high affinity is in urgent need to develop immunoassay for screening purpose. In this work, an unreported hapten, 5-(propylthio)-1H-benzo[d]imidazol-2-amine, was designed and synthesized, which maximally exposed the characteristic sulfanyl group of ABZ to the animal immune system to induce expected antibody. One monoclonal antibody (Mab) that can simultaneously detect ABZs was obtained with IC50 values of 0.20, 0.26, 0.77, and 10.5 μg/L for ABZ, ABZSO2, ABZSO, and ABZNH2SO2 in ic-ELISA under optimized conditions respectively, which has been never achieved in previous reports. For insight into the recognition profiles of the Mab, we used computational chemistry method to parameterize cross-reactive molecules in aspects of conformation, electrostatic fields, and hydrophobicity, revealing that the hydrophobicity and conformation of characteristic group of molecules might be the key factors that together influence antibody recognition with analytes. Furthermore, the practicability of the developed ic-ELISA was verified by detecting ABZs in spiked milk, beef, and liver samples with recoveries of 60% to 108.8% and coefficient of variation (CV) of 1.0% to 15.9%.

ESTIMATION OF PERIODONTAL MICROCIRCULATION, JAW AFTER INJECTION OF ANTI-RESORPTIVE DRUG IN EXPERIMENT

Importance. Determining the influence of different medications in the development of different diseases is a prerequisite for adequate comprehensive treatment of patients. At present, there is insufficient data on the influence of anti-osteoclastic drugs on the microcirculation, which may have an impact. The aim. To study the effects of the monoclonal antibody denosumab on the development of osteonecrosis of the jaw of rats. Methodology. The study was carried out on 36 Wistar Line rats in the Department of Pathophysiology with the course of clinical pathophysiology First Pavlov State Medical University. The osteonecrorosis of the jaws of rats was induced by the extracted of a lower first molar. The observation was carried out with diagnostic studies of microcirculation (doppler), bone structure (3D computer tomography of jaws), followed by statistical processing of the data. Results. It has been shown that the greatest decrease in blood flow rate and the greatest bone defect is determined in the group of rats with the maximum dose of the monoclonal antibody preparation of denosumab by the time of 4 weeks, 0.5 mg/kg intravenous administration. Also, there has been evidence of a difference in blood flow reduction between the histological layers of the lower jaw. Thus, the intrabone blood flow was damaged much more, than the layer of mucous membrane of the gum. Conclusions. The combined method of dopplerography allows the determination of blood flow status on different histological layers. It is possible to determine the extent to which different pharmacological preparations influence the rate of blood flow in the local area without invasive interventions. The study of monoclonal antibodies is an acute problem in the world of surgical dental practice, which requires further study.

Application of Microfluidic Technology in Field of Antibody Preparation

Microfluidic technology is a science and technology that can accurately manipulate fluids in micro-sized channels. In recent years, microfluidic devices have attracted wide attention due to its easy manipulation, miniaturized size, high throughput and precise control, which provide a potential platform for antibody screening. This review paper provides an overview of recent advances in microfluidic methods application in the field of antibody preparation. While hybridoma technology and four antibody engineering techniques including phage display, single B cell antibody screening, antibody expression and cell-free protein synthesis are mainly introduced, important advances of experimental models and results are also discussed. Furthermore, the authors expound on the limitations of current microfluidic screening system and present future directions of antibody screening platform based on microfluidics. Antibody preparation on microfluidics combined with other technologies has huge application potential in the field of biomedicine, and it is anticipated to be further developed.

Evaluation of liquid and powdered forms of polyclonal antibody preparation against Streptococcus bovis and Fusobacterium necrophorum in cattle adapted or not adapted to highly fermentable carbohydrate diets

Objective: Feed additives that modify rumen fermentation can be used to prevent metabolic disturbances such as acidosis and optimize beef cattle production. The study evaluated the effects of liquid and powdered forms of polyclonal antibody preparation (PAP) against <i>Streptococcus bovis</i> and Fusobacterium necrophorum on rumen fermentation parameters in ruminally cannulated non-lactating dairy cows that were adapted or unadapted to a high concentrate diet.Methods: A double 3×3 Latin square design was used with three PAP treatments (control, powdered, and liquid PAP) and two adaptation protocols (adapted, unadapted; applied to the square). Adapted animals were transitioned for 2 weeks from an all-forage to an 80% concentrate diet, while unadapted animals were switched abruptly.Results: Interactions between sampling time and adaptation were observed; 12 h after feeding, the adapted group had lower ruminal pH and greater total short chain fatty acid concentrations than the unadapted group, while the opposite was observed after 24 h. Acetate:propionate ratio, molar proportion of butyrate and ammonia nitrogen concentration were generally greater in adapted than unadapted cattle up to 36 h after feeding. Adaptation promoted 3.5 times the number of <i>Entodinium</i> protozoa but copy numbers of <i>Streptococcus bovis</i> and Fibrobacter succinogens genes in rumen fluid were not affected. However, neither liquid nor powdered forms of PAP altered rumen acidosis variables in adapted or unadapted animals.Conclusion: Adaptation of cattle to highly fermentable carbohydrate diets promoted a more stable ruminal environment, but PAP was not effective in this study in which no animal experienced acute or sub-acute rumen acidosis.

124 Polyclonal Antibody Preparations from Avian Origin Increased Fiber Digestibility of Beef Cattle Receiving a Backgrounding Diet

This study investigated the effects of feeding an avian-derived polyclonal antibody preparation (PAP; CAMAS, Inc.) against Streptococcus bovis, Fusobacterium necrophorum, and lipopolysaccharides (40, 35, and 25% of the preparation, respectively) on plasmatic haptoglobin and glucose concentrations (Exp. 1), and apparent total tract digestibility (Exp. 2) of beef cattle consuming a backgrounding diet. In Exp. 1, Angus crossbreed heifers and steers (n = 90; 373 ± 62 kg BW) were randomly assigned to receive a common ad libitum diet (76% TDN, 15.9% CP, DM basis) with the addition of 1 (PAP1), 3 (PAP3), or 0 g (CON) of PAP daily. Plasmatic concentrations of glucose and haptoglobin were measured on d 0, 14, 28, 42, and 56. In Exp. 2, 25 Angus crossbreed steers (390 ± 65 kg BW) were used in a completely randomized design. Steers were housed in 3 pens to receive the same diet and treatments from Exp. 1. Feed intake was measured using GrowSafe feed bunks, and indigestible neutral detergent fiber was used as the internal marker. In Exp. 1, a day effect was detected for plasmatic haptoglobin and glucose concentrations (P ≤ 0.04). In Exp. 2, no difference in DMI was observed (P = 0.88). Dry matter, organic matter, and acid detergent fiber digestibility in the total tract were reduced (P ≤ 0.05), whereas CP digestibility tended to decrease (P ≤ 0.07) in steers receiving PAP3 vs. CON and PAP1. Apparent total tract starch digestibility was increased (P ≤ 0.02) for PAP1 vs. PAP3. Neutral detergent fiber digestibility was greater (P &lt; 0.01) for PAP1 vs. CON and PAP3. Feeding 1 g/d of a PAP against Streptococcus bovis, Fusobacterium necrophorum, and lipopolysaccharides increased fiber digestibility in backgrounding diets; however, further research is needed to understand the impaired responses on nutrient digestibility when greater doses are provided.

123 Polyclonal antibody preparations from avian origin did not reduce plasmatic haptoglobin concentrations but increased ruminal NH3-N in beef steers during transition from forage to high-grain diets

This study investigated the effects of feeding an avian-derived polyclonal antibody preparation (PAP; CAMAS, Inc.) against Streptococcus bovis, Fusobacterium necrophorum, and lipopolysaccharides (40, 35, and 25% of the preparation, respectively) on plasmatic haptoglobin, ruminal short chain fatty acids (SCFA), and ammonia-N (NH3-N) concentrations of beef steers during a 21-d step-up adaptation to a high-grain diet. Eight ruminally cannulated Angus crossbred beef steers (658 ± 79 kg BW) were randomly assigned to treatments in a crossover design to be transitioned from a diet containing bermudagrass hay [Cynodon dactylon (L.) Pers.] plus 0.45 kg/d of molasses with 0 (CON) or 3 g/d of PAP (PAP) to a high-grain diet. Transition consisted of three 7-d steps of increased inclusion of cracked corn (35, 60, and 80% diet DM for STEP1, STEP2, and STEP3, respectively). On each transition d and 7 d after STEP3 (STEP3-7d), ruminal fluid samples were obtained every 3 h for 24 h. Blood samples were collected on d 0, 1, and 3 relative to each transition for haptoglobin determinations. Haptoglobin plasmatic concentrations increased (P = 0.03) on d 2 and 3 vs. d 1 during STEP2 and on STEP3 compared to STEP1 and STEP2 (P = 0.01). Steers receiving PAP had greater (P &lt; 0.01) ruminal NH3-N concentration in STEP1; however, there were no effects of treatment on SCFA (P &gt; 0.10). Total SCFA concentrations were affected by the step-up diets (P &lt; 0.01); propionate concentration (Pro) was greater in STEP2 through STEP3-7d vs. STEP1 (P &lt; 0.01), whereas acetate concentration (Ac) and Ac:Pro linearly decreased from STEP1 to STEP3-7d (P &lt; 0.01). Feeding 3 g/d of polyclonal antibody preparations against Streptococcus bovis, Fusobacterium necrophorum, and lipopolysaccharides in a 21-d step-up adaptation to high-grain diets did not affect plasmatic haptoglobin or ruminal SCFA concentrations; however, it increased ruminal NH3-3 concentrations.

Advances in Antibody Preparation Techniques for Immunoassays of Total Aflatoxin in Food

Aflatoxin (AF) contamination is a major concern in the food and feed industry because of its prevalence and toxicity. Improved aflatoxin detection methods are still needed. Immunoassays are an important method for total aflatoxin (TAF) analysis in food due to its technical advantages such as high specificity, sensitivity, and simplicity, but require high-quality antibodies. Here, we first review the three ways to prepare high-quality antibodies for TAF immunoassay, second, compare the advantages and disadvantages of antigen synthesis methods for B-group and G-group aflatoxins, and third, describe the status of novel genetic engineering antibodies. This review can provide new methods and ideas for the development of TAF immunoassays.

A Mechanism of Gold Nanoparticle Aggregation by Immunoglobulin G Preparation

Conjugates of gold nanoparticles (GNPs) and antibodies are widely used in various fields of biochemistry and microbiology. However, the procedure for obtaining such conjugates remains precarious, and the properties of conjugates differ significantly for different antibody clones. One of the most common problems is the aggregation of GNPs in the course of their conjugation with antibodies. This article considers an example of the conjugation of monoclonal antibodies with non-stable aggregating product. The composition of the antibody preparation was studied using electrophoresis, asymmetrical flow field-flow fractionation, and ultracentrifugation. It was shown that the component that causes the aggregation of the GNPs is the light chains of immunoglobulins that appear due to the spontaneous decay of the antibodies. After separation of the fraction with a molecular weight of less than 30 kDa, stable conjugates of antibodies with GNPs were obtained. The high functional activity of the obtained conjugates was confirmed by immunochromatography.