Publications by authors named "Kenneth W Lee"

20 Publications

  • Page 1 of 1

Multivariable analysis of the influence of cross-reactive carbohydrate determinant inhibition and other factors on intradermal and serological allergen test results: a prospective, multicentre study.

Vet Dermatol 2021 Jun 9. Epub 2021 Jun 9.

Department of Clinical Sciences and Advanced Medicine, School of Veterinary Medicine, University of Pennsylvania, 3900 Spruce Street, Philadelphia, PA, 19104, USA.

Background: Serological allergen testing (SAT) is used widely to formulate allergen-specific immunotherapy for atopic dogs. Serum immunoglobulin (Ig)E specific for cross-reactive carbohydrate determinants (CCD) can produce false-positive reactions, creating discrepancy between SAT and intradermal allergen test (IDAT) results.

Objectives: The primary objective was to determine if inhibition of anti-CCD IgE in a commercial assay improved correlation with IDAT. The secondary objective was to assess the influence of dog- and clinic-specific factors, environmental factors, putative allergen exposure and prior medications on intradermal and SAT reactivity.

Animals: Two-hundred and eleven client-owned dogs were enrolled from eight North American dermatology specialty practices.

Methods And Materials: Collection of serum samples and IDAT were performed on the same day. Sera were assayed for detection of IgE specific to 25 allergens, before and after treatment with a proprietary inhibitor of anti-CCD IgE. Data for each dog were collected via a questionnaire filled out by veterinary personnel.

Results: The correlation between the testing modalities was fair before (Spearman's rho, ρ = 0.2092) and after (ρ = 0.3042) inhibition of anti-CCD IgE. Ciclosporin dose (P = 0.003), independent of duration of use, and duration of lokivetmab use (P = 0.001), independent of dose administered, were associated with statistically significant decreases in IgE concentrations across all allergen types.

Conclusions And Clinical Relevance: Contrary to previous reports, this study demonstrated unchanged correlation between SAT and IDAT after inhibition of anti-CCD IgE. Ciclosporin dose and lokivetmab treatment duration may have unexplored effects on IgE concentration during SAT.
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http://dx.doi.org/10.1111/vde.12974DOI Listing
June 2021

Detection and inhibition of IgE antibodies reactive with cross-reactive carbohydrate determinants in an ELISA for allergen-specific IgE in horses.

Vet Dermatol 2021 May 6. Epub 2021 May 6.

Montesano & Tallarico, LLP, PO Box 1396, Smithtown, NY, USA.

Background: It has been demonstrated that immunoglobulin (Ig)E specific for cross-reactive carbohydrate determinants (CCD) is present in the serum of sensitized humans, dogs and cats, and that these CCD-specific antibodies might confound serological testing.

Hypothesis/objective: The objective was to determine whether or not CCD-reactive antibodies occur in horses and to investigate the prevalence of CCD-reactive IgE antibodies in equine sera using a monoclonal cocktail-based enzyme-linked immunosorbent assay designed to detect allergen-specific IgE in horses, and to evaluate a means for successful inhibition of these CCD.

Methods And Materials: Sera from 28 horses suspected of clinical allergy were evaluated, with and without a proprietary inhibitor which contains carbohydrates derived from bromelain (BROM-CCD), using a panel of 72 allergens that include 15 grasses, 17 trees, nine weeds, five mites, 12 fungi, 12 insects and two environmental allergens.

Results: Twenty-five samples were shown to be reactive to at least one of the allergens, and 15 were reactive to 10 allergens or more. BROM-CCD had minimal effect on the mite reactivity in any of the positive samples; however, substantial inhibition for pollen allergens (trees, grasses and weeds) was demonstrable. Reduction in signal to pollens ranged from 20% to 100% for samples that were inhibited by CCD-BROM.

Conclusions And Clinical Importance: These results demonstrate that CCD-reactive IgE antibodies are evident in horses and that BROM-CCD can be effective in reducing reactions with these irrelevant carbohydrates and will likely yield a more accurate in vitro allergen reactivity profile for selection of allergens included in an immunotherapeutic regime.
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http://dx.doi.org/10.1111/vde.12963DOI Listing
May 2021

Mass Analysis of Macro-molecular Analytes via Multiply-Charged Ion Attachment.

Anal Chem 2020 12 4;92(24):16301-16306. Epub 2020 Dec 4.

Department of Chemistry, Purdue University, 560 Oval Drive, West Lafayette, Indiana 47907-2084, United States.

A novel gas-phase charge and mass manipulation approach is demonstrated to facilitate the mass measurement of high mass complexes within the context of native mass spectrometry. Electrospray ionization applied to solutions generated under native or near-native conditions has been demonstrated to be capable of preserving biologically relevant complexes into the gas phase as multiply charged ions suitable for mass spectrometric analysis. However, charge state distributions tend to be narrow and extensive salt adduction, heterogeneity, and so on tend to lead to significantly broadened peaks. These issues can compromise mass measurement of high mass bio-complexes, particularly when charge states are not clearly resolved. In this work, we show that the attachment of high mass ions of known mass and charge to populations of ions of interest can lead to well-separated signals that can yield confident charge state and mass assignments from otherwise poorly resolved signals.
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http://dx.doi.org/10.1021/acs.analchem.0c04335DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7738406PMC
December 2020

Digital ion trap mass analysis of high mass protein complexes using IR activation coupled with ion/ion reactions.

Int J Mass Spectrom 2020 Dec 20;458. Epub 2020 Sep 20.

Department of Chemistry, Purdue University, West Lafayette, IN, 47907-2084, USA.

Native mass spectrometry (MS) focuses on measuring the masses of large biomolecular complexes and probing their structures. Large biomolecular complexes are readily introduced into mass spectrometers as gas-phase ions using electrospray ionization (ESI); however, the ions tend to be heavily adducted with solvent and salts, which leads to mass measurement errors. Various solution clean-up approaches can reduce the degree of adduction prior to introduction to the mass spectrometer. Gas-phase activation of trapped ions can provide additional adduct reduction, and charge reduction ion/ion reactions increase charge state separation. Together, gas-phase activation and charge reduction can combine to yield spectra of well separated charge states for improved mass measurements. A simple gas-phase collisional activation technique is to apply a dipolar DC (DDC) field to opposing electrodes in an ion trap. DDC activation loses its efficacy when ions are trapped at low values, which is true of the high ions generated by charge reduction ion/ion reactions. Digital ion trapping (DIT) readily traps high ions at higher values by varying trapping frequency rather than amplitude, but the low frequencies used to trap high ions also decreases the efficacy of DDC activation. We demonstrate here using ions derived from GroEL that IR activation of ions shows no discrimination against high ions trapped with DIT, because they can be focused equally well to the trap center to interact with the IR laser beam. Following pump out of excess background gas, IR activation can also induce efficient dissociation of the GroEL complex. This work demonstrates that IR activation is an effective approach for ion heating in native MS over the unusually wide range of charge states accessible via gas-phase ion/ion reactions.
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http://dx.doi.org/10.1016/j.ijms.2020.116437DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7641502PMC
December 2020

Detection and Inhibition of IgE for cross-reactive carbohydrate determinants evident in an enzyme-linked immunosorbent assay for detection of allergen-specific IgE in the sera of dogs and cats.

Vet Dermatol 2020 Dec 25;31(6):439-e116. Epub 2020 Sep 25.

School of Veterinary Medicine, University of Pennsylvania, 3900 Spruce St, Philadelphia, PA, 19104, USA.

Background: It has been demonstrated recently that immunoglobulin (Ig)E specific for cross-reactive carbohydrate determinants (CCD) is present in the serum of allergen-sensitized dogs and cats, and that these CCD-specific antibodies might confound serological testing.

Hypothesis/objective: The objective was to document the prevalence of CCD detectable in a monoclonal cocktail-based enzyme-linked immunosorbent assay designed for the detection of allergen-specific IgE in the sera of dogs and cats, and to define a means for successful inhibition of these CCD.

Methods And Materials: The incidence of reactivity to bromelain and a commercially available inhibitor of carbohydrate-specific antibodies (RIDA-CCD) was evaluated in 100 dog sera samples before and after inhibition with RIDA-CCD and a proprietary inhibitor containing carbohydrates derived from bromelain (BROM-CCD). Subsequently, sera from 600 dogs and 600 cats were evaluated using a serum diluent with and without BROM-CCD.

Results: Both the RIDA-CCD and BROM-CCD inhibitors demonstrated successful reduction of CCD reactivity, although a more efficient profile of inhibition was evident with BROM-CCD. Mite reactivity in dog and cat sera was largely unaffected; however, substantial inhibition for pollen allergens (trees, grasses and weeds) was shown. After BROM-CCD inhibition, 1% of canine samples and 13% of feline samples were rendered completely negative for allergen reactivity.

Conclusions And Clinical Importance: The results demonstrate that BROM-CCD is effective in reducing reactions with irrelevant carbohydrates, and that inhibition of CCD reactivity might substantially alter the outcome of the in vitro reactivity profile used for selection of allergens to be included in an immunotherapeutic regime.
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http://dx.doi.org/10.1111/vde.12904DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7756665PMC
December 2020

Ion trap operational modes for ion/ion reactions yielding high mass-to-charge product ions.

Int J Mass Spectrom 2020 May 21;451. Epub 2020 Feb 21.

Department of Chemistry, Purdue University, West Lafayette, IN, 47907-2084, USA.

To better probe large biomolecular complexes, developments in mass spectrometry (MS) have focused on improving technologies used to generate, transmit, and measure high ions. The additional tandem-MS (MS) capabilities of ion trap mass spectrometers (ITMS) facilitate experiments that facilitate probing complex biomolecular ions. In particular, charge reduction using gas-phase ion/ion reactions increase separation of charge states generated via electrospray ionization (ESI), which increases confidence in charge state assignments and therefore masses determined from the observed charge states. Current ITMS technologies struggle to generate and measure low charge states of large (>50 kDa) proteins and complexes because of power limitations associated with conventional high-frequency sine wave operation. Other approaches, including frequency scanning techniques and use of digital waveforms, reduce or eliminate some of these limitations. The work presented here studies five different operational modes for a quadrupole ion trap (QIT) mass spectrometer used to generate and measure low charge states of bovine serum albumin (BSA), pyruvate kinase (PK), and GroEL. While digital operation of a QIT presents limitations during the ion/ion reaction period of the experiment, it generally provided the best spectra in terms of resolution and signal at > 50,000.
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http://dx.doi.org/10.1016/j.ijms.2020.116313DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7189770PMC
May 2020

Valet Parking for Protein Ion Charge State Concentration: Ion/Molecule Reactions in Linear Ion Traps.

Anal Chem 2020 04 9;92(7):5419-5425. Epub 2020 Mar 9.

Department of Chemistry, Purdue University, West Lafayette, Indiana 47907-2084, United States.

There are several analytical applications in which it is desirable to concentrate analyte ions generated over a range of charge states into a single charge state. This has been demonstrated in the gas phase via ion/ion reactions in conjunction with a technique termed ion parking, which can be implemented in electrodynamic ion traps. Ion parking depends upon the selective inhibition of the reaction of a selected charge state or charge states. In this work, we demonstrate a similar charge state concentration effect using ion/molecule reactions rather than ion/ion reactions. The rates of ion/molecule reactions cannot be affected in the manner used in conventional ion parking. Rather, to inhibit the progression of ion/molecule proton transfer reactions, the product ions must be removed from the reaction cell as they are formed and transferred to an ion trap where no reactions occur. This is accomplished here with mass-selective axial ejection (MSAE) from one linear ion trap to another. The application of MSAE to inhibit ion/molecule reactions is referred to as "valet parking" as it entails the transport of the ions of interest to a remote location for storage. Valet parking is demonstrated using model proteins to concentrate ion signal dispersed over multiple charge states into largely one charge state. Additionally, it has been applied to a simple two-protein mixture of cytochrome and myoglobin.
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http://dx.doi.org/10.1021/acs.analchem.0c00146DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7145756PMC
April 2020

Increasing the Upper Mass/Charge Limit of a Quadrupole Ion Trap for Ion/Ion Reaction Product Analysis via Waveform Switching.

J Am Soc Mass Spectrom 2019 Jun 14;30(6):1126-1132. Epub 2019 Mar 14.

Department of Chemistry, Purdue University, 560 Oval Drive, West Lafayette, IN, 47907-2084, USA.

Quadrupole ion traps (QITs) are versatile platforms for performing experiments with gas-phase ions due to their abilities to store ions of both polarities and to conduct MS experiments. The QIT is particularly useful as a reaction cell for ion/ion reactions. In the case of an ion/ion reaction experiment in a QIT, multiply charged reactant ions may initially be of relatively low m/z (e.g., m/z < 1000) whereas the product ions can be one or more orders of magnitude higher in m/z (e.g., m/z > 100,000). Several factors can limit the m/z range over which an ion/ion reaction experiment can be conducted. These include (1) the efficiency of the detector, (2) the m/z range over which oppositely charged ions can be mutually stored, and (3) the m/z range over which ions can be mass selectively ejected into an external detector. High-frequency waveforms provide larger m/z trapping ranges for mutual storage of oppositely charged ions whereas low-frequency waveforms provide better trapping for very high m/z product ions. Presented here is a method that switches from a high-frequency sine wave prior to and during an ion/ion reaction to a low-frequency square wave to eject low m/z reagent ions and improves confinement of the product ions before mass-selective ejection by scanning the frequency of the square wave. This approach addresses the third issue, which is the primary limiting factor with QITs operated at high RF (e.g., > 900 MHz). Graphical Abstract.
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http://dx.doi.org/10.1007/s13361-019-02156-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6520138PMC
June 2019

Longitudinal evaluation of immunological responses to allergen-specific immunotherapy in horses with IgE associated dermatological disease, a pilot study.

Vet Dermatol 2019 Jun 4;30(3):255-e78. Epub 2019 Mar 4.

Greer Laboratories, Inc, 639 Nuway Circle NE, Lenoir, NC, 28645, USA.

Background: The long-term effects of allergen specific immunotherapy (ASIT) on concentrations of circulating immunoglobulin E (IgE) and immunoglobulin G (IgG) in horses have not been reported.

Objectives: To document changes in clinical severity of horses with atopic dermatitis (AD) and to monitor allergen-specific IgE and IgG concentrations during a two-year course of ASIT.

Animals: Nineteen client-owned horses with a conditional diagnosis of AD.

Methods And Materials: Three ASIT groups were randomly assigned based upon results obtained by either intradermal testing (IDT) for regional allergens (n = 7); enzyme-linked immunosorbent assay (ELISA) for specific IgE (n = 6); or a composite of results from both tests (n = 6). Serum concentrations of IgE and IgG specific for allergens included in ASIT were measured at time zero and at four-month intervals. A visual analog scale (VAS) was used to record severity of clinical signs at times zero, 12 and 24 months.

Results: Positive correlations were documented between IgE and both immediate and delayed IDT results (P < 0.00001), and between immediate IDT and IgG results (P = 0.003). Specific IgE in sera decreased significantly (P < 0.05) for allergens that were included in ASIT, whereas IgG increased. Across all horses, the mean VAS score decreased by 1.2 units [95% CI: 1.28, 1.14; (P < 0.0001)] during each 12-month period of ASIT therapy. Improvement in clinical signs was noted in 76.5% of the horses following 12 months of ASIT and in 82% after 24 months on ASIT.

Conclusions And Clinical Importance: In this pilot study, ASIT in horses with AD provided significant clinical benefit associated with a concomitant reduction of allergen-specific IgE and elevation of IgG.
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http://dx.doi.org/10.1111/vde.12732DOI Listing
June 2019

A Miniaturized Fourier Transform Electrostatic Linear Ion Trap Mass Spectrometer: Mass Range and Resolution.

J Am Soc Mass Spectrom 2019 Apr 12;30(4):588-594. Epub 2019 Feb 12.

Department of Chemistry, Purdue University, 560 Oval Drive, West Lafayette, IN, 47907-2084, USA.

Mass resolution (M/ΔM) was increased by reducing the axial length of a Fourier transform electrostatic linear ion trap (FT-ELIT) mass spectrometer. The increase in mass resolution corresponds directly to increased axial ion frequencies in the FT-ELIT. Increased mass resolution was demonstrated for equivalent transient lengths in a 5.25″ versus 2.625″ ELIT using the isotopes of [bradykinin+2H] and [insulin+5H] as test ions. Both bradykinin and insulin show mass resolution increases of ~ 90% allowing baseline resolution of the [insulin+5H] isotopes after only 300 ms of data acquisition. Relative changes in mass/charge range were explored using mirror switching to trap ions injected axially into the ELIT. When trapping ions using mirror switching, the mass/charge range in a FT-ELIT mass spectrometer for a given switch time is determined by the time required for fast ions to enter and exit the trap after one reflection versus the time it takes for slow ions to enter the trap. By reducing the length of the FT-ELIT mass spectrometer while maintaining a constant distance from the point from which ions are initially accelerated to the entrance mirror, only the low m/z limit is affected for a given mirror switching time. For the two ELIT lengths examined here, the effective mass/charge range at any given switch time is reduced from m/z-8.9*m/z for the 5.25″ ELIT to m/z-5.2*m/z for the 2.625″ ELIT. Graphical Abstract.
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http://dx.doi.org/10.1007/s13361-018-02126-xDOI Listing
April 2019

Fourier-Transform MS and Closed-Path Multireflection Time-of-Flight MS Using an Electrostatic Linear Ion Trap.

Anal Chem 2017 10 3;89(20):10965-10972. Epub 2017 Oct 3.

Department of Chemistry Purdue University West Lafayette, Indiana 47907-2084, United States.

An electrostatic linear ion trap (ELIT) has been configured to allow for the simultaneous acquisition of mass spectra via Fourier transform (FT) techniques (frequency measurement) and via time-of-flight (TOF; time measurement). In the former case, the time-domain image charge derived from a pick-up electrode in the field-free region of the ELIT is converted to frequency-domain data via Fourier transformation (i.e., FT-ELIT MS). In the latter case, the time difference between ion injection into the ELIT and ion detection after release from the ELIT using a microchannel plate (MCP) enables the acquisition of multireflection time-of-flight mass spectra (MR-TOF MS). The ELIT geometry facilitates the acquisition of both types of data simultaneously because the detection schemes are independent and do not preclude one another. The two MS approaches exhibit a degree of complementarity. Resolution increases much faster with time with the MR-TOF approach, for example, but the closed-path nature of executing MR-TOF in an ELIT limits both the m/z range and the peak capacity. For this reason, the FT-ELIT MS approach is most appropriate for wide m/z range applications, whereas MR-TOF MS can provide advantages in a "zoom-in" mode in which moderate resolution (M/ΔM ≈ 10000) at short analysis times (10 ms) is desirable.
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http://dx.doi.org/10.1021/acs.analchem.7b02797DOI Listing
October 2017

Determination of Collision Cross Sections Using a Fourier Transform Electrostatic Linear Ion Trap Mass Spectrometer.

J Am Soc Mass Spectrom 2018 02 11;29(2):242-250. Epub 2017 Jul 11.

Department of Chemistry, Purdue University, 560 Oval Drive, West Lafayette, IN, 47907-2084, USA.

Collision cross sections (CCSs) were determined from the frequency-domain linewidths in a Fourier transform electrostatic linear ion trap. With use of an ultrahigh-vacuum precision leak valve and nitrogen gas, transients were recorded as the background pressure in the mass analyzer chamber was varied between 4× 10 and 7 × 10 Torr. The energetic hard-sphere ion-neutral collision model, described by Xu and coworkers, was used to relate the recorded image charge to the CCS of the molecule. In lieu of our monoisotopically isolating the mass of interest, the known relative isotopic abundances were programmed into the Lorentzian fitting algorithm such that the linewidth was extracted from a sum of Lorentzians. Although this works only if the isotopic distribution is known a priori, it prevents ion loss, preserves the high signal-to-noise ratio, and minimizes the experimental error on our homebuilt instrument. Six tetraalkylammonium cations were used to correlate the CCS measured in the electrostatic linear ion trap with that measured by drift-tube ion mobility spectrometry, for which there was an excellent correlation (R ≈ 0.9999). Although the absolute CCSs derived with our method differ from those reported, the extracted linear correlation can be used to correct the raw CCS. With use of [angiotensin II] and reserpine, the corrected CCSs (334.9 ± 2.1 and 250.1 ± 0.5, respectively) were in good agreement with the reported ion mobility spectrometry CCSs (335 and 254.3, respectively). With sufficient signal-to-noise ratio, the CCSs determined are reproducible to within a fraction of a percent, comparable to the uncertainties reported on dedicated ion mobility instruments. Graphical Abstract ᅟ.
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http://dx.doi.org/10.1007/s13361-017-1720-1DOI Listing
February 2018

Engineering Surfaces through Sequential Stop-Flow Photopatterning.

Adv Mater 2016 Nov 12;28(42):9292-9300. Epub 2016 Sep 12.

Materials Research Laboratory (MRL), University of California, Santa Barbara, Santa Barbara, CA, 93106, USA.

Solution-exchange lithography is a new modular approach to engineer surfaces via sequential photopatterning. An array of lenses reduces features on an inkjet-printed photomask and reproduces arbitrarily complex patterns onto surfaces. In situ exchange of solutions allows successive photochemical reactions without moving the substrate and affords access to hierarchically patterned substrates.
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http://dx.doi.org/10.1002/adma.201602900DOI Listing
November 2016

Ear Mite Removal in the Santa Catalina Island Fox (Urocyon littoralis catalinae): Controlling Risk Factors for Cancer Development.

PLoS One 2015 7;10(12):e0144271. Epub 2015 Dec 7.

Karen C. Drayer Wildlife Health Center, School of Veterinary Medicine, University of California Davis, Davis, California, United States of America.

Ear mites (Otodectes cynotis) and ear canal tumors are highly prevalent among federally endangered Island foxes (Urocyon littoralis catalinae) living on Santa Catalina Island off the coast of Southern California. Since studies began in the 1990s, nearly all foxes examined were found to be infected with ear mites, and ceruminous gland tumors (carcinomas and adenomas) were detected in approximately half of all foxes ≥ 4 years of age. We hypothesized that reduction of ear mite infection would reduce otitis externa and ceruminous gland hyperplasia, a risk factor for tumor development. In this study, we conducted a randomized field trial to assess the impact of acaricide treatment on ear mite prevalence and intensity of infection, otitis externa, ceruminous gland hyperplasia, and mite-specific IgG and IgE antibody levels. Treatment was highly effective at eliminating mites and reducing otitis externa and ceruminous gland hyperplasia, and mite-specific IgG antibody levels were significantly lower among uninfected foxes. Ceruminous gland hyperplasia increased in the chronically infected, untreated foxes during the six month study. Our results provide compelling evidence that acaricide treatment is an effective means of reducing ear mites, and that mite removal in turn reduces ear lesions and mite-specific IgG antibody levels in Santa Catalina Island foxes. This study has advanced our understanding of the underlying pathogenesis which results in ceruminous gland tumors, and has helped inform management decisions that impact species conservation.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0144271PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4671584PMC
June 2016

Proficiency monitoring of monoclonal antibody cocktail-based enzyme-linked immunosorbent assay for detection of allergen-specific immunoglobulin E in dogs.

J Vet Diagn Invest 2015 Jul 11;27(4):461-9. Epub 2015 Jun 11.

Greer Laboratories Inc., Lenoir, NC (Lee, Blankenship, McKinney)IDEXX GmbH, Ludwigsburg, Germany (Kern)IDEXX Laboratories Inc., Westbrook, ME (Buch)IDEXX Laboratories Inc. Markham, Ontario, Canada (Greenwood)Univet Diagnostic Services, Sant Cugat, Barcelona, Spain (Brazis)Biovac, Beaucouzé Cedex, France (Drouet)Agrolabo SpA, Scarmagno, Torino, Italy (Tambone)Artu Biologicals, Almere, The Netherlands (Faas)Axiom Veterinary Laboratories, Newton Abbot, Devon, UK (Weaver).

The purpose of our study was to document the continued comparative proficiency of different laboratories that perform a monoclonal antibody-based enzyme-linked immunosorbent assay (macELISA) for detection of allergen-specific immunoglobulin (Ig)E in dogs. Replicate samples of 18 different sera pools were independently evaluated in a single blinded fashion by each of 16 different operators functioning in 10 different laboratories. The average intra-assay variance among reactive assay calibrators in all laboratories was 6.0% (range: 2.7-16.1%), while the average intralaboratory interassay variance was 7.5% (range: 3.9-10.9%). The overall interassay interlaboratory variance was consistent among laboratories and averaged 11.4% (range: 8.5-12.5%). All laboratories yielded similar profiles and magnitudes of responses for replicate unknown samples; dose response profiles observed in each of the laboratories were indistinguishable. Considering the positive or negative results, interassay interlaboratory concordance of results exceeded 90%. Correlation of optical density values between and among all laboratories was strong (r > 0.9, P < 0.001). Collectively, the results demonstrated that the macELISA for measuring allergen-specific canine IgE is reproducible, and documents that consistency of results can be achieved not only in an individual laboratory by differing operators but also among laboratories using the same monoclonal-based ELISA.
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http://dx.doi.org/10.1177/1040638715587547DOI Listing
July 2015

Dynamic strain-mediated coupling of a single diamond spin to a mechanical resonator.

Nat Commun 2014 Jul 18;5:4429. Epub 2014 Jul 18.

Department of Physics, University of California Santa Barbara, Broida Hall, Santa Barbara, California 93106, USA.

The development of hybrid quantum systems is central to the advancement of emerging quantum technologies, including quantum information science and quantum-assisted sensing. The recent demonstration of high-quality single-crystal diamond resonators has led to significant interest in a hybrid system consisting of nitrogen-vacancy centre spins that interact with the resonant phonon modes of a macroscopic mechanical resonator through crystal strain. However, the nitrogen-vacancy spin-strain interaction has not been well characterized. Here, we demonstrate dynamic, strain-mediated coupling of the mechanical motion of a diamond cantilever to the spin of an embedded nitrogen-vacancy centre. Via quantum control of the spin, we quantitatively characterize the axial and transverse strain sensitivities of the nitrogen-vacancy ground-state spin. The nitrogen-vacancy centre is an atomic scale sensor and we demonstrate spin-based strain imaging with a strain sensitivity of 3 × 10(-6) strain Hz(-1/2). Finally, we show how this spin-resonator system could enable coherent spin-phonon interactions in the quantum regime.
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http://dx.doi.org/10.1038/ncomms5429DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4109012PMC
July 2014

Intra and inter-laboratory reproducibility of a monoclonal antibody cocktail based ELISA for detection of allergen specific IgE in dogs: proficiency monitoring of macELISA in six laboratories.

Vet Immunol Immunopathol 2012 Aug 23;148(3-4):267-75. Epub 2012 May 23.

Greer Laboratories Inc., 639 Nuway Circle, Lenoir, NC 28645, USA.

The purpose of this study was to evaluate the reproducibility of results yielded using a monoclonal antibody based ELISA for detection of allergen specific IgE when run in six separate affiliated laboratories. On two separate occasions, duplicate samples of 15 different sera pools were independently evaluated by each laboratory in a single blinded fashion. The average intra-assay variance among reactive assay calibrators in all laboratories was 6.2% (range 2.6-18.2%), while the average intra-laboratory inter-assay variance was 12.1% (range 8.0-17.1%). The overall inter-assay inter-laboratory variance was consistent among laboratories and averaged 15.6% (range 15.1-16.6%). All laboratories yielded similar profiles and magnitudes of responses for replicate unknown samples; dose-response profiles observed in each of the laboratories were indistinguishable. Considering positive/negative results, inter-assay inter-laboratory concordance of results exceeded 95%. Correlation of OD values between and among all laboratories was strong (r>0.9, p<0.001). Correlation of OD values between the two separate evaluations was also high for all allergens except olive, which was attributed to lot-to-lot differences of allergen coated wells. Collectively, the results demonstrated that the monoclonal antibody based ELISA for measuring allergen specific canine IgE is reproducible, and documents that consistency of results can be achieved not only in an individual laboratory, but between laboratories using the same monoclonal-based ELISA.
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http://dx.doi.org/10.1016/j.vetimm.2012.05.011DOI Listing
August 2012

Performance characteristics of a monoclonal antibody cocktail-based ELISA for detection of allergen-specific IgE in dogs and comparison with a high affinity IgE receptor-based ELISA.

Vet Dermatol 2009 Jun 3;20(3):157-64. Epub 2009 Apr 3.

Greer Laboratories, Inc., Lenoir, North Carolina 28645, USA.

The purpose of this study was to define the operational and performance characteristics of a commercially available monoclonal antibody based (mac) ELISA for detection of allergen-specific IgE in dogs. The average intra-assay variance over 1 year was 9.7% (range 2.5-62.7%), while the interassay variance averaged 10.8% (range 8.1-13.8%). The average positive control responses observed for grass, weed, tree and mite allergens during each month remained relatively constant; the average monthly variance was 11.6% (range 8.3-19.2%) for grass pollens, 13.3% (range 9.1-20.4%) for weed pollens, 13.3% (range 9.8-18.2%) for tree pollens and 13.6% (range 8.9-18.7%) for mite allergens. The interlaboratory concordance of results for the macELISA was approximately 91%. The interlaboratory concordance of results comparing the macELISA and a high affinity IgE receptor-based ELISA was approximately 92%. The results demonstrate that the macELISA is reproducible and the results are comparable to the high affinity IgE receptor based ELISA within and between laboratories.
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http://dx.doi.org/10.1111/j.1365-3164.2009.00740.xDOI Listing
June 2009

Assessment of cross-reactivity among five species of house dust and storage mites.

Vet Dermatol 2008 Apr;19(2):67-76

Clinic of Medicine, School of Veterinary Medicine, University of Thessaly, Karditsa, Greece.

In vitro cross-reactivity among two house dust (Dermatophagoides farinae, D. pteronyssinus) and three storage (Acarus siro, Tyrophagus putrescentiae, Lepidoglyphus destructor) mites was examined in 20 mite-sensitive dogs with natural occurring atopic dermatitis (group A), 13 high-IgE beagles experimentally sensitized to D. farinae (group B), and five healthy beagles (group C). Intradermal testing (IDT) and serology for allergen-specific IgE demonstrated that co-sensitization for all possible pairs of the five mites was generally 45% or higher among group A dogs. In the same dogs, enzyme-linked immunosorbent assay cross-inhibition results indicated that each one of D. farinae, A. siro and T. putrescentiae was a strong inhibitor of all the remaining mites, whereas D. pteronyssinus was a strong inhibitor of L. destructor. A high number of positive IDT and serology test results for D. pteronyssinus, A. siro, T. putrescentiae and L. destructor were recorded among group B dogs. No conclusive evidence of exposure to these mites was found upon analysis of dust samples from their environment and their food for the presence of mites and guanine. Also, the number of positive test results was generally higher among group B than among group C dogs. Enzyme-linked immunosorbent assay cross-inhibition revealed that D. farinae was a strong inhibitor of D. pteronyssinus, A. siro and T. putrescentiae. Collectively, these results demonstrated extensive in vitro cross-reactivity among house dust and/or storage mites that can explain false-positive results upon testing of dust mite-sensitive dogs with atopic dermatitis.
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http://dx.doi.org/10.1111/j.1365-3164.2008.00654.xDOI Listing
April 2008

The immunopathogenesis of flea allergy dermatitis in dogs, an experimental study.

Vet Immunol Immunopathol 2004 Jun;99(3-4):179-92

Department of Diagnostic Medicine/Pathobiology, College of Veterinary Medicine, Kansas State University, Manhattan, KS 66506, USA.

In this study, we investigated the development of clinical disease and immune responses in the development of an experimental model of flea allergy dermatitis. Dogs were randomly divided into four treatment groups and were infested with fleas on two different feeding schedules (continuous and episodic). Group 1 consisted of four non-exposed dogs (negative controls) and Group 2 consisted of six dogs exposed to fleas continually. Groups 3 and 4 consisted of 14 dogs each that were exposed to fleas on an episodic schedule (two consecutive days every other week for 12 weeks). Group 4 also received intraperitoneal injections of a low dose of lectin (ricin) with immunomodulatory properties. The purpose of Group 4 was to investigate the effects of ricin on enhancing the development of clinical signs, flea antigen-specific IgE levels and altering the number of CD4+ and CD8+ T cell subsets in peripheral blood. Clinical signs developed in all flea exposed dogs, however, the dermatology lesion scores were less and shorter in duration for continuously exposed dogs compared to episodic exposed dogs, independent of ricin treatment. Lesion development was concentrated in the flea triangle and consisted principally of erythema, followed by alopecia, excoriation, papules, and crusts. CD4+ and CD8+ lymphocyte subsets or IgE levels were not altered by ricin treatment. Flea antigen-specific IgE values were highest in dogs exposed to fleas on a continuous basis compared to those episodically exposed. A greater percentage of clinical responder dogs with negative flea-specific IgE titers or negative intradermal test (IDT) were present in the episodic exposure groups than in the continuous exposure group. IgE titers corresponded slightly better with clinical responders than the IDT. The agreement between the IgE titers and IDT was good (weighted K = 0.67). Histopathology of skin samples were consistent with a Type I hypersensitivity. In conclusion, we were able to develop a model of flea allergy dermatitis by experimentally exposing dogs to fleas on an episodic and continuous feeding schedule. In this study, continuously exposed dogs did not develop immunotolerance, and ricin did not enhance the development of FAD.
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http://dx.doi.org/10.1016/j.vetimm.2004.02.006DOI Listing
June 2004