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Kuck L.R.,Indevr, Inc. | Saye S.,Indevr, Inc. | Loob S.,Indevr, Inc. | Roth-Eichhorn S.,Glaxosmithkline | And 2 more authors.
Vaccine | Year: 2017

Vaccine manufacturers require more rapid and accurate tools to characterize the potency and stability of their products. Currently, the gold standard for influenza vaccine potency is the single radial immunodiffusion (SRD) assay, which has inherent disadvantages. The primary objective of this study was to investigate the ability of the VaxArray Influenza (VXI) seasonal hemagglutinin (sHA) potency assay to accurately quantify potency and stability in finished vaccines as well as to quantify hemagglutinin protein (HA) within crude in-process samples. Monobulk intermediates and mono- and multivalent vaccines were tested using VXI. Quantification of HA in crude samples was evaluated by spiking known concentrations of HA into allantoic fluid. VXI generated SRD equivalent potency measurements with high accuracy (within ±10%) and precision (CV 10 ± 4%) for antigen components of monobulk intermediates and multivalent split vaccines. For these vaccines and vaccine intermediates, the VXI linear dynamic range was ∼0.01–0.6 μg/mL, which is 12× greater than the linear range of SRD. The measured sample limit of detection (LOD) for VXI varied from 0.005 to 0.01 μg/mL for the different subtypes, which in general is ≥600× lower than the LOD for SRD. VXI was able to quantify HA in crude samples where HA only accounts for 0.02% of the total protein content. Stability indication was investigated by tracking measured potency as a function of time at elevated temperature by both SRD and VXI. After 20 h at 56 °C, the ratio of VXI to SRD measured potency in a quadrivalent vaccine was 76%, 125%, 60%, and 98% for H1/California, H3/Switzerland, B/Phuket and B/Brisbane, respectively. Based on the study results, it is concluded that VXI is a rapid, multiplexed immunoassay that can be used to accurately determine flu vaccine potency and stability in finished product and in crude samples from upstream processes. © 2017 The Author(s)


Wilson G.,Indevr, Inc. | Ye Z.,U.S. Food and Drug Administration | Xie H.,U.S. Food and Drug Administration | Vahl S.,Indevr, Inc. | And 2 more authors.
PLoS ONE | Year: 2017

The hemagglutination inhibition assay (HAI) is widely used to evaluate vaccine-induced antibody responses as well as to antigenically characterize influenza viruses. The results of an HAI assay are based on an endpoint titration where the titers are generally manually interpreted and recorded by a well-trained expert. For serological applications, the lack of standardization in endpoint interpretation and interference from non-specific inhibitors in clinical samples can translate into a high degree of variability in the results. For example, tilting HAI plates at 45±60 degrees to look for a 'tear drop pattern' with avian red blood cells is a common practice by many, but not all, research laboratories. In this work, we tested the hypothesis that an automated image analysis algorithm can be used to achieve an accurate and non-subjective interpretation of HAI assays-specifically without the need to tilt plates. In a side-by-side comparison study performed during FDA's biannual serological screening process for influenza viruses, titer calls for more than 2200 serum samples were made by the Cypher One automated hemagglutination analyzer without tilting and by an expert human with tilting. The comparison yielded 95.6% agreement between the expert reader and automated interpretation method (within ± 1 dilution) for the complete dataset. Performance was also evaluated relative to the type of red blood cell (Turkey and Guinea pig) and influenza strain (12 different viruses). For the subset that utilized Guinea pig red blood cells (~44% of the samples), for which no plate tilting was required, the agreement with an expert reader was 97.2%. For the subset that utilized Turkey red blood cells (~56% of the samples), for which plate tilting was necessary by the expert reader, the agreement was 94.3%. Overall these results support the postulate that algorithm-based interpretation of a digital record with no plate tilting could replace manual reading for greater consistency in HAI assays. © This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.


Heil G.L.,University of Florida | McCarthy T.,University of Iowa | Yoon K.-J.,Iowa State University | Liu S.,Iowa State University | And 6 more authors.
Influenza and other Respiratory Viruses | Year: 2010

Please cite this paper as: Heil et al. (2010) MChip, a low density microarray, differentiates among seasonal human H1N1, North American swine H1N1, and the 2009 pandemic H1N1. Influenza and Other Respiratory Viruses 4(6), 411-416. Background The MChip uses data from the hybridization of amplified viral RNA to 15 distinct oligonucleotides that target the influenza A matrix (M) gene segment. An artificial neural network (ANN) automates the interpretation of subtle differences in fluorescence intensity patterns from the microarray. The complete process from clinical specimen to identification including amplification of viral RNA can be completed in <8hours for under US$10. Objectives The work presented here represents an effort to expand and test the capabilities of the MChip to differentiate influenza A/H1N1 of various species origin. Methods The MChip ANN was trained to recognize fluorescence image patterns of a variety of known influenza A viruses, including examples of human H1N1, human H3N2, swine H1N1, 2009 pandemic influenza A H1N1, and a wide variety of avian, equine, canine, and swine influenza viruses. Robustness of the MChip ANN was evaluated using 296 blinded isolates. Results Training of the ANN was expanded by the addition of 71 well-characterized influenza A isolates and yielded relatively high accuracy (little misclassification) in distinguishing unique H1N1 strains: nine human A/H1N1 (88·9% correct), 35 human A/H3N2 (97·1% correct), 31 North American swine A/H1N1 (80·6% correct), 14 2009 pandemic A/H1N1 (87·7% correct), and 23 negative samples (91·3% correct). Genetic diversity among the swine H1N1 isolates may have contributed to the lower success rate for these viruses. Conclusions The current study demonstrates the MChip has the capability to differentiate the genetic variations among influenza viruses with appropriate ANN training. Further selective enrichment of the ANN will improve its ability to rapidly and reliably characterize influenza viruses of unknown origin. © 2010 Blackwell Publishing Ltd.


Quinones B.,U.S. Department of Agriculture | Swimley M.S.,U.S. Department of Agriculture | Taylor A.W.,Indevr, Inc. | Dawson E.D.,Indevr, Inc.
Foodborne Pathogens and Disease | Year: 2011

Shiga toxin-producing Escherichia coli O157 is a leading cause of foodborne illness worldwide. To evaluate better methods to rapidly detect and genotype E. coli O157 strains, the present study evaluated the use of ampliPHOX, a novel colorimetric detection method based on photopolymerization, for pathogen identification with DNA microarrays. A low-density DNA oligonucleotide microarray was designed to target stx1 and stx2 genes encoding Shiga toxin production, the eae gene coding for adherence membrane protein, and the per gene encoding the O157-antigen perosamine synthetase. Results from the validation experiments demonstrated that the use of ampliPHOX allowed the accurate genotyping of the tested E. coli strains, and positive hybridization signals were observed for only probes targeting virulence genes present in the reference strains. Quantification showed that the average signal-to-noise ratio values ranged from 47.73 ± 7.12 to 76.71 ± 8.33, whereas average signal-to-noise ratio values below 2.5 were determined for probes where no polymer was formed due to lack of specific hybridization. Sensitivity tests demonstrated that the sensitivity threshold for E. coli O157 detection was 100-1000 CFU/mL. Thus, the use of DNA microarrays in combination with photopolymerization allowed the rapid and accurate genotyping of E. coli O157 strains. © 2011, Mary Ann Liebert, Inc.


Vajzovic A.,University of Washington | Bura R.,University of Washington | Kohlmeier K.,Indevr, Inc. | Doty S.L.,University of Washington
Journal of Industrial Microbiology and Biotechnology | Year: 2012

A systematic study was conducted characterizing the effect of furfural, 5-hydroxymethylfurfural (5-HMF), and acetic acid concentration on the production of xylitol and ethanol by a novel endophytic yeast, Rhodotorula mucilaginosa strain PTD3. The influence of different inhibitor concentrations on the growth and fermentation abilities of PTD3 cultivated in synthetic nutrient media containing 30 g/l xylose or glucose were measured during liquid batch cultures. Concentrations of up to 5 g/l of furfural stimulated production of xylitol to 77 % of theoretical yield (10 % higher compared to the control) by PTD3. Xylitol yields produced by this yeast were not affected in the presence of 5-HMF at concentrations of up to 3 g/l. At higher concentrations of furfural and 5-HMF, xylitol and ethanol yields were negatively affected. The higher the concentration of acetic acid present in a media, the higher the ethanol yield approaching 99 % of theoretical yield (15 % higher compared to the control) was produced by the yeast. At all concentrations of acetic acid tested, xylitol yield was lowered. PTD3 was capable of metabolizing concentrations of 5, 15, and 5 g/l of furfural, 5-HMF, and acetic acid, respectively. This yeast would be a potent candidate for the bioconversion of lignocellulosic sugars to biochemicals given that in the presence of low concentrations of inhibitors, its xylitol and ethanol yields are stimulated, and it is capable of metabolizing pretreatment degradation products. © Society for Industrial Microbiology and Biotechnology 2012.


Kuck L.R.,Indevr, Inc. | Sorensen M.,Protein Sciences Corporation | Matthews E.,Protein Sciences Corporation | Srivastava I.,Protein Sciences Corporation | And 2 more authors.
PLoS ONE | Year: 2014

Titer on Chip (Flu-ToC) is a new technique for quantification of influenza hemagglutinin (HA) concentration. In order to evaluate the potential of this new technique, a comparison of Flu-ToC to more conventional methods was conducted using recombinant HA produced in a baculovirus expression system as a test case. Samples from current vaccine strains were collected from four different steps in the manufacturing process. A total of 19 samples were analysed by Flu-ToC (blinded), single radial immunodiffusion (SRID), an enzyme-linked immunosorbent assay (ELISA), and the purity adjusted bicinchoninic acid assay (paBCA). The results indicated reasonable linear correlation between Flu-ToC and SRID, ELISA, and paBCA, with regression slopes of log-log plots being 0.91, 1.03, and 0.91, respectively. The average ratio for HA content measured by Flu-ToC relative to SRID, ELISA, and paBCA was 83%, 147%, and 81%, respectively; indicating nearly equivalent potency determination for Flu-ToC relative to SRID and paBCA. These results, combined with demonstrated multiplexed analysis of all components within a quadrivalent formulation and robust response to HA strains over a wide time period, support the conclusion that Flu-ToC can be used as a reliable and time-saving alternative potency assay for influenza vaccines. © 2014 Kuck et al.


PubMed | Protein Sciences Corporation and Indevr, Inc.
Type: Journal Article | Journal: PloS one | Year: 2014

Titer on Chip (Flu-ToC) is a new technique for quantification of influenza hemagglutinin (HA) concentration. In order to evaluate the potential of this new technique, a comparison of Flu-ToC to more conventional methods was conducted using recombinant HA produced in a baculovirus expression system as a test case. Samples from current vaccine strains were collected from four different steps in the manufacturing process. A total of 19 samples were analysed by Flu-ToC (blinded), single radial immunodiffusion (SRID), an enzyme-linked immunosorbent assay (ELISA), and the purity adjusted bicinchoninic acid assay (paBCA). The results indicated reasonable linear correlation between Flu-ToC and SRID, ELISA, and paBCA, with regression slopes of log-log plots being 0.91, 1.03, and 0.91, respectively. The average ratio for HA content measured by Flu-ToC relative to SRID, ELISA, and paBCA was 83%, 147%, and 81%, respectively; indicating nearly equivalent potency determination for Flu-ToC relative to SRID and paBCA. These results, combined with demonstrated multiplexed analysis of all components within a quadrivalent formulation and robust response to HA strains over a wide time period, support the conclusion that Flu-ToC can be used as a reliable and time-saving alternative potency assay for influenza vaccines.


Morfa A.J.,National Renewable Energy Laboratory | Barnes T.M.,National Renewable Energy Laboratory | Ferguson A.J.,National Renewable Energy Laboratory | Levi D.H.,National Renewable Energy Laboratory | And 3 more authors.
Journal of Polymer Science, Part B: Polymer Physics | Year: 2011

We describe a comprehensive model for the optical properties of pristine films of poly-(3-hexylthiophene) (P3HT). The presented model is anisotropic with the optical axis normal to the substrate plane, which is consistent with previous x-ray diffraction studies that show preferential edge-on packing of the polymer chains on the substrate. Peak locations and spacings are defined using a Huang-Rhys vibronic progression consistent with known phonon energies. We demonstrate that the model fits variable-angle spectroscopic ellipsometry and normal-incidence transmission data well, and accurately predicts angle- and polarization-dependent transmission and reflection data. The spectral features of the optical constants used in the model are in excellent agreement with published spectroscopic data on P3HT. © 2010 Wiley Periodicals, Inc.


Indevr, Inc. | Entity website

InDevR leads way at IQ Innovation Awards with two victories BOULDER Life science company InDevR cleaned up at the 15th edition of BizWests Innovation Quotient Awards, taking home the awards for Innovative Company of the Year and the peoples choice award for Innovation of the Year. The IQ Awards were held at the Boulder Theater, and featured 21 innovative companies from the Boulder Valley pitching in seven different products and services categories ...


PubMed | Indevr, Inc.
Type: Journal Article | Journal: Vaccine | Year: 2015

Readers of this Special Issue of Vaccine are likely unified by a desire to enhance vaccine efficacy and improve vaccine manufacturing efficiency. For influenza vaccines, challenges to achieving those goals are many, and range from improved surveillance to less cumbersome production platforms and more reliable performance verification. Specifically, demand is growing for an alternative vaccine potency assay. Assuming that a replacement potency assay is found to be promising, one question will be: how will it be judged to be accurate? It is generally agreed that any potential replacement for SRID will have to exhibit good correlation with SRID and yield a value within 20% of the SRID determined potency. In my opinion, SRID itself has enough limitations that judging alternatives relative to that particular standard will not ensure that the industry as a whole transitions to an improved method. In fact, it could blind us to an assay that may ultimately provide a better predictive measure of vaccine efficacy (or immunogenicity). There is only one way to verify that measured potency is correlated with, and predictive of, measured immunogenicity - test the relationship in clinical studies.

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