Factors influencing the economic mechanism of the development of small and medium-sized dairy cattle breeding enterprises

This article includes analysis of factors influencing economic mechanism for the development of small and medium-sized enterprises in the area of dairy farming. To accomplish the main task, the authors used economic parameters of the production results of small and medium-sized enterprises according to the financial statements of federal and regional government bodies, as well as official data from the Central Bank of Russia, the Territorial Authority of the Federal State Statistics Service for the Chelyabinsk Region and data from the Ministry of Agriculture and Federal Tax Service. The Chelyabinsk Region was chosen as a typical region for study. This research was conducted based on data from the register of small and medium-sized enterprises. Based on statistical data, small and medium-sized enterprises were divided into groups, economic parameters were calculated for each group of subjects, quantitative factors were defined that have the greatest impact on the efficiency of the economic mechanism for the development of small and medium-sized enterprises

Simultaneous measurements of auto-immune and infectious disease specific antibodies using a high throughput multiplexing tool.

Considering importance of ganglioside antibodies as biomarkers in various immune-mediated neuropathies and neurological disorders, we developed a high throughput multiplexing tool for the assessment of gangliosides-specific antibodies based on Biolpex/Luminex platform. In this report, we demonstrate that the ganglioside high throughput multiplexing tool is robust, highly specific and demonstrating ∼100-fold higher concentration sensitivity for IgG detection than ELISA. In addition to the ganglioside-coated array, the high throughput multiplexing tool contains beads coated with influenza hemagglutinins derived from H1N1 A/Brisbane/59/07 and H1N1 A/California/07/09 strains. Influenza beads provided an added advantage of simultaneous detection of ganglioside- and influenza-specific antibodies, a capacity important for the assay of both infectious antigen-specific and autoimmune antibodies following vaccination or disease. Taken together, these results support the potential adoption of the ganglioside high throughput multiplexing tool for measuring ganglioside antibodies in various neuropathic and neurological disorders

Testing multiplexing capacity of combined ganglioside-influenza bead array.

<p>Luminex beads coated with various payloads of Brisbane H1N1 and California H1N1 hemagglutinins were tested, and beads with the payload 0.5 µg (encircled in red) was selected for the combined ganglioside-influenza bead array (a, b). As a proof of principle, ganglioside-influenza bead array was tested with sera from two donors vaccinated against seasonal influenza in 2009 (c) and against pandemic California H1N1 in 2010 (d). Increase of influenza-specific antibodies post vaccination is apparent. More details in the text. (Note: Errors were too small to include in the panels a and b). Anti-Brisbane H1N1 and anti-California H1N1 sheep sera (NIBSC, UK) were used at dilution 1∶ 50000 as a positive control for detecting influenza-specific antibodies by hemagglutinin-coated beads. Anti-ganglioside rabbit sera (Matreya LLC) were used as positive controls for detecting ganglioside-specific antibodies at the following dilutions: anti-GM1 - 1∶6400, anti-GM2 - 1∶800, anti-GA1 – 1∶25000, anti-GD1B – 1∶400.</p

Concentration sensitivity to ganglioside-specific sera of BioPlex bead array, compared to ELISA.

<p>Ganglioside-conjugated Luminex bead exhibited approximately 100 times better sensitivity to the sera specific to GM1, GM2 and GG1b gangliosides (a, b, d), and about 10 times better sensitivity to the serum specific to GA1 (c). Both BioPlex and ELISA assays used the same detecting antibody, anti-rabbit IgG(H+L)Fab2:biotin at 2 ug/ml, and SA-PE fluorescent tag at 4 ug/ml. Dashed circles show approximate sensitivity thresholds for BioPlex (blue) and ELISA (red).</p

Cross-reactivity of ganglioside antibodies.

<p>GM1- and GA1-conjugated beads showed considerable cross reactivity with anti-GD1b sera since these gangliosides have common terminal sugar sequence <b>(panels b, c; dotted red square in panel e)</b>. GM1 beads displayed cross-reactivity with anti-GM2 serum due to the shared epitope groups in the inner part of the GM1- and GM2-gangliosides <b>(panel d; dotted black square in panel e)</b>. Cross-reactivity was estimated comparing the reporter signal from the beads coated with the ganglioside related to the tested serum and the signals from the beads coated with other gangliosides.</p

Evaluating bead epitope specificity using CTB and AS epitope blockers.

<p>CTB has ability to bind 5 GM1 molecules (<b>a</b>) compared to alpha-synuclein that can bind only one GM1 molecule (<b>b</b>). CTB showed ∼90% bead blocking with anti-GM1 rabbit serum compared to ∼15% with other anti-ganglioside rabbit sera while synuclein exhibits ∼60–90% blocking with all anti-ganglioside sera (<b>c</b>). Screening human sera, 20%–40% blocking was seen with CTB compared to 70%–80% using AS (<b>d</b>). The radical R circled in dashed red signifies the presence of sialic acid groups (one in GK1 and GM2, two in GD1b and none in GA1).</p

Fluorescence Adherence Inhibition Assay: A Novel Functional Assessment of Blocking Virus Attachment by Vaccine-Induced Antibodies.

Neutralizing antibodies induced by vaccination or natural infection play a critically important role in protection against the viral diseases. In general, neutralization of the viral infection occurs via two major pathways: pre- and post-attachment modes, the first being the most important for such infections as influenza and polio, the latter being significant for filoviruses. Neutralizing capacity of antibodies is typically evaluated by virus neutralization assays that assess reduction of viral infectivity to the target cells in the presence of functional antibodies. Plaque reduction neutralization test, microneutralization and immunofluorescent assays are often used as gold standard virus neutralization assays. However, these methods are associated with several important prerequisites such as use of live virus requiring safety precautions, tedious evaluation procedure and long assessment time. Hence, there is a need for a robust, inexpensive high throughput functional assay that can be performed rapidly using inactivated virus, without extensive safety precautions. Herein, we report a novel high throughput Fluorescence Adherence Inhibition assay (fADI) using inactivated virus labeled with fluorescent secondary antibodies virus and Vero cells or erythrocytes as targets. It requires only few hours to assess pre-attachment neutralizing capacity of donor sera. fADI assay was tested successfully on donors immunized with polio, yellow fever and influenza vaccines. To further simplify and improve the throughput of the assay, we have developed a mathematical approach for calculating the 50% titers from a single sample dilution, without the need to analyze multi-point titration curves. Assessment of pre- and post-vaccination human sera from subjects immunized with IPOL®, YF-VAX® and 2013-2014 Fluzone® vaccines demonstrated high efficiency of the assay. The results correlated very well with microneutralization assay performed independently by the FDA Center of Biologics Evaluation and Research, with plaque reduction neutralization test performed by Focus Diagnostics, and with hemaglutination inhibition assay performed in-house at Sanofi Pasteur. Taken together, fADI assay appears to be a useful high throughput functional immunoassay for assessment of antibody-related neutralization of the viral infections for which pre-attachment neutralization pathway is predominant, such as polio, influenza, yellow fever and dengue

fADI screening of pre- and post-vaccinated serum samples from donors immunized with IPOL^® vaccine.

<p>The screening was performed in the single dilution mode rather than using sera titration. Donor-to-donor variation in the capacity of donor sera to block virus attachment was observed for all three poliovirus serotypes (A, B and C). The control ELISA tests (D, E and F) did not display significant reduction of the reporter fluorescence signal, thus showing very low or zero competition between test serum antibodies and virus-specific labeling antibodies used in the assay.</p