Genotyping of clinically relevant human adenoviruses by array-in-well hybridization assay

AbstractA robust oligonucleotide array-in-well hybridization assay using novel up-converting phosphor reporter technology was applied for genotyping clinically relevant human adenovirus types. A total of 231 adenovirus-positive respiratory, ocular swab, stool and other specimens from 219 patients collected between April 2010 and April 2011 were included in the study. After a real-time PCR amplification targeting the adenovirus hexon gene, the array-in-well assay identified the presence of B03 (n = 122; 57.5% of patients), E04 (29; 13.7%), C02 (21; 9.9%), D37 (14; 6.6%), C01 (12; 5.7%), C05 (5; 2.4%), D19 (4; 1.9%), C06 (2; 0.9%), D08 (1; 0.5%), A31 (1; 0.5%) and F41 (1; 0.5%) genotypes among the clinical sample panel. The typing result was obtained for all specimens that could be amplified (n = 223; 97%), and specificity of the typing was confirmed by sequencing specimens representing each of the different genotypes. No hybridization signal was obtained in adenovirus-negative specimens or specimens with other viruses (n = 30). The array-in-well hybridization assay has great potential as a rapid and multiplex platform for the typing of clinically relevant human adenovirus genotypes in different specimen types

Lapin platinaryhmän mineraalihippujen koostumus ja synty

Tiivistelmä. Platinaryhmän mineraalihippuja (platinum group elements minerals (PGM)) saadaan sivutuotteena kullankaivuun yhteydessä Lapin kullanhuuhdonta-alueilta. Tärkein hyödynnetty mineraalia on isoferroplatina. Hippuja esiintyy granuliittikaaren sisällä olevissa upaesiintymissä ja niiden lähtöpaikka on tuntematon. Tutkielman tarkoituksena on selvittää hipuissa esiintyvien PGM-sulkeumien mineralogiaa ja tehdä johtopäätöksiä niiden muodostumisesta. Sulkeumien mineralogiaa ja koostumusta vertaillaan maailmalla esiintyviin platinaryhmän alkuaine-esiintymiin. Tutkimusta varten hankittiin näytteitä lapin kullanhuuhdonta-alueilta. Näytteitä saatiin Lemmenjoen- Ivalojoen ja Tankavaara-Mäkärä-Roivainen alueilta. Näytteistä valmistettiin kiillotetut pintahieet. Kaikkiaan 50 platinaryhmän mineraaliraetta tutkittiin FESEM pyyhkäisyelektronimikroskoopin avulla. Sulkeumat analysoitiin laitteen energiadispersiivisellä analysaattorilla (EDS) ja tulkinta suoritettiin laskemalla analyysistä mineraalin ionisuhteet. Analyysejä otettiin kaikkiaan 1000:sta sulkeumasta. Tuloksista havainnoitiin 22 eri PGM-mineraalia ja seosta. Yleisimpinä sulkeumamineraaleina esiintyi Ru-Os-disulfidit lauriiti-erlichmaniitti, jotka muodostavat vyöhykkeellisiä sulkeumia isoferroplatinan sisään. Seuraavaksi yleisimmät mineraalit olivat PGE-sulfarseniitit, joista tärkein on Irarsiitti. Lisäksi esiintyi rodarseniittia, hollingwortiittia. Ru-Os-Ir-seokset esiintyivät sulkeumina rakeiden sisustassa, joista osa muodosti kritallografisesti suuntautuneita suotaumia. Tiospinelli ryhmästä esiintyi cuproiridsiitti-malaniitti. Pt-Pd-Au-Cu-seoksia esiintyi omina rakeinaan. Pt-Fe-Cu-seokset esiintyivät sulkeumina isoferroplatinan sisässä ja reunassa, lisäksi yhdessä rakeessa esiintyi nimeämätön Pt2Cu-mineraali hongshiniitin ympäröimänä. Perusmetallisulfideja esiintyi sulkeumana yhdessä rakeessa. Osa makroskooppisesti tulkituista isoferroplatinahipuista paljastui Au-Hg- ja weishaniittihipuiksi. Tutkimuksessa löydetyistä mineraaleista Os-Ir-Ru-seokset, lauriitti-erlichmaniiti ovat muodostuneet magmaattisesti korkeassa lämpötilassa. Lämpötilan laskiessa muodostui cuproiridsiitti, kashiniitti sulkeumat(suotaumat?). Myöhäismagmaattisessa ja hydrotermisessä tapahtumassa kehittyi PGE-sulfarsenidit, atokiitti, cooperiitti ja speryllitti sulkeumat sekä osa Pt-Fe-Cu seoksista. Yksi Pt-Fe-Cu-rae osoittaa magmaatisen kiteytymisen piirteitä. Vyöhykkeellinen Pd-Pt-rae voi rakenteensa puolesta supergeenisesti muodostunut. Tutkimus osoitti, että isoferroplatinahiput voivat olla peräisin ofioliittisesta, Ural-Alaska -tyypin intruusiosta tai kerrosintruusiosta. Ruteniumpitoiset rakeet indikoivat ofioliittista lähtöainesta ja sulfidiköyhät puolestaan Ural-Alaska tyypin esiintymää. Sperylliitti rakeet voivat olla peräisin Ni-Cu-PGE-esiintymästä tai kerrosintruusiosta. Suotuisimmat kivilajit esiintymille sijaitsevat Tenojoki-vyöhykeessä granuliittikaaren ulkopuolella. Aiemmin Lapista ei ole ilmoitettu Au-Hg-hippuja. Rakeiden mineralogia osoittaa toisenlaista lähtöpaikkaa hipuille kuin yleisesti oletetut kvartsi-karbonaatti juonet.The composition and origin of the platinum group mineral nuggets in the Finnish Lapland. Abstract. Platinum group mineral (PGM) nuggets have been escavated from goldpanning areas as a side product. Most important PGM’s are isoferroplatinum nuggets. The aim of this work is to analyse inclusion from isoferroplatinum and make conclusions concerning their genesis. The mineralogy and compositions of the inclusion will be compared with platinum group element (PGE) deposits worldwide. Samples were obtained from Lemmenjoki, Ivalojoki and Tankavaara-Mäkärä-Roivainen gold panning areas in Lapland. Polished sections were prepared from the samples. A total of 50 platinum group mineral grains were examined by Zeiss Ultra Plus field emission scanning electron microscope (FESEM). The analyses were carried out on the energy dispersive spectrometer (EDS) of the instrument and the interpretation was performed by calculating the ionic ratios of the mineral from the analysis. A total of 1000 inclusions were analysed. Twenty two different PGM minerals and alloys were found in the samples. The most common inclusions were Ru-Os disulphides laurite and erlichmanite, which form zonal inclusions within the isoproteroplatinum. The next most common minerals were PGE-sulfarsenides with irarsite being the most common. Rhodarsenide, hollingworthite occur as a smaller inculsion in the rutenium alloys and irarsite. Ru-Os-Ir alloys occur as small inclusions in the grain interiors, some of which formed crystallographically oriented exsolutions. From the thiospinelli group, cuproiridsite-malanite occurs in grain boundaries and as inclusions. Pt-Pd-Au-Cu mixtures appeared as their own grains. Pt-Fe-Cu mixtures appeared as inclusions and at the margin of isoferroplatinum, in addition, one grain contained an unnamed Pt2Cu mineral surrounded by hongshinite. Basemetal sulphides cubanite, pyrrhotite and chalcopyrite were present in one grain. Some of the macroscopically interpreted isoferroplatinum nuggets were revealed as Au-Hg and weishanite nuggets. The Os-Ir-Ru alloys, laurite-erichmanite minerals were formed magmatically at high temperature. As the temperature decreased, cuproiridsite, kashinite exolutions formed. In a late magmatic and hydrothermal event, PGE-sulfarsenides, atokite, cooperite and speryllite inclusions developed. One Pt-Fe-Cu grain shows features of magmatic crystallization. The study showed that isoferroplatinum nuggets may be derived from Ophiolitic source, Ural-Alaska type intrusions or Layered Intrusions. The ruthenium-rich grains indicate an Ophiolite source and the sulphide-poor ones a Ural-Alaska type intrusion. The source of speryllite grains may be Ni-Cu-PGE deposit or a layered intrusion. The most favourable rock types for the deposits are located in the Tenojoki-belt outside the granulite belt. In the past, no Au-Hg nuggets have been reported from Lapland. The mineralogy of the nuggets indicates a different origin for the grains than the commonly assumed quartz-carbonate veins

Upconversion Cross-Correlation Spectroscopy of a Sandwich Immunoassay

Fluorescence correlation and cross-correlation spectroscopy (FCS/FCCS) have enabled biologists to study processes of transport, binding, and enzymatic reactions in living cells. However, applying FCS and FCCS to samples such as whole blood and plasma is complicated as the fluorescence bursts of diffusing labels can be swamped by strong autofluorescence. Here we present cross-correlation spectroscopy based on two upconversion nanoparticles emitting at different wavelengths on the anti-Stokes side of a single excitation laser. This upconversion cross-correlation spectroscopy (UCCS) approach allows us to completely remove all Stokes shifted autofluorescence background in biological material such as plasma. As a proof of concept, we evaluate the applicability of UCCS to a homogeneous sandwich immunoassay for thyroid stimulating hormone measured in buffer solution and in plasma

Gingival cell growth with antiresorptive treatment combined with corticosteroids or antiestrogen

Objectives: Antiresorptive treatment has been shown to impair mucosal cell proliferation, migration, and viability. However, in the clinic, antiresorptives are often used in combination with other drugs. We studied the effect of antiresorptives combined with a corticosteroid or antiestrogen on oral mucosal keratinocytes and fibroblasts.Material and methods: Human gingival keratinocyte and fibroblast cell lines were exposed to bisphosphonates (BPs) and denosumab in different concentrations and durations together with an antiestrogen or corticosteroid. Changes in cell viability, proliferation and migration after exposures were measured. Data were evaluated with hierarchical linear mixed model for repeated measurements.Results: Bisphosphonate exposure suppressed keratinocyte and fibroblast cell viability, proliferation, and migration in a time-dependent manner. Combining a corticosteroid or antiestrogen with BPs further increased this negative effect. Denosumab alone had a mild positive effect on keratinocyte and fibroblast growth. When denosumab was combined with a corticosteroid or antiestrogen, cell growth was suppressed.Conclusions: Our results show that coexisting medications may increase the negative impact of BPs or denosumab on oral mucosal cells

A randomised clinical study to determine the effect of a toothpaste containing enzymes and proteins on plaque oral microbiome ecology

The numerous species that make up the oral microbiome are now understood to play a key role in establishment and maintenance of oral health. The ability to taxonomically identify community members at the species level is important to elucidating its diversity and association to health and disease. We report the overall ecological effects of using a toothpaste containing enzymes and proteins compared to a control toothpaste on the plaque microbiome. The results reported here demonstrate that a toothpaste containing enzymes and proteins can augment natural salivary defences to promote an overall community shift resulting in an increase in bacteria associated with gum health and a concomitant decrease in those associated with periodontal disease. Statistical analysis shows significant increases in 12 taxa associated with gum health including Neisseria spp. and a significant decrease in 10 taxa associated with periodontal disease including Treponema spp. The results demonstrate that a toothpaste containing enzymes and proteins can significantly shift the ecology of the oral microbiome (at species level) resulting in a community with a stronger association to health

Effective Shielding of NaYF4:Yb3+,Er3+ Upconverting Nanoparticles in Aqueous Environments Using Layer-by-Layer Assembly

Aqueous solutions are the basis for most, biomedical assays, but they quench the upconversion luminescence significantly. Surface modifications of upconverting nanoparticles are vital for shielding the obtained luminescence. Modifications also provide new possibilities for further use by introducing attaching sites for biomolecule conjugation. We demonstrate the use of a layer-by-layer surface modification method combining varying lengths of negatively charged polyelectrolytes with positive neodymium ions in coating the upconverting NaYF4:Yb3+,Er3+ nanoparticles. We confirmed the formation of the bilayers and investigated the surface properties with Fourier transform infrared and reflectance spectroscopy, thermal analysis, and (zeta-potential measurements. The effect of the coating on the upconversion luminescence properties was characterized, and the bilayers with the highest improvement in emission intensity were identified. In addition, studies for the nanoparticle and surface stability were carried out in aqueous environments. It was observed that the bilayers were able to shield the materials' luminescence from quenching also in the presence of phosphate buffer that is currently considered the most disruptive environment for the nanoparticles

Nanoparticle-aided glycovariant assays to bridge biomarker performance and ctDNA results

AbstractNumerous immunoassay based cancer biomarkers established in the 1970 and 1980'ies are widely used in clinical routine. Initial expectations of biomarkers such as CEA, CA125, CA19-9, AFP to provide decisive help in the diagnosis of early stage, pre-symptomatic cancers have not been realized. Thus, they are primarily used for monitoring disease progression and occasionally being useful as prognostic indicators. This limitation is due to the marker also being measurable in healthy individuals and frequently at elevated concentrations in common benign conditions. Most conventional tumor markers are glycosylated and interestingly specific alterations of the glycostructure part can often be seen early in the cancerous process. Conventional double monoclonal immunoassays are however blind to such changes as they are based on peptide epitope recognition. Wide selections of carbohydrate recognizing macromolecules, lectins, but also glycan structure recognizing antibodies are potentially useful for detecting such changes. Despite numerous attempts generating proof-of-principle evidence for this, such assays have generally not been successfully introduced into clinical routine. The affinity constants of lectin and glycan specific antibodies for their corresponding carbohydrate structures may be up to several orders too low to provide the detection limits and robustness expected from routine tumor markers. In this review, we describe an approach based on the use of highly fluorescent Eu3+--chelate dyed nanoparticles onto which lectins or glycan specific antibodies are coated to provide the necessary binding strength and signal amplification to provide low detection limits, while maintaining the original glycan-structure specificity. This concept applied to three markers, PSA, CA125 and CA15-3 provide glycoform assays of greatly enhanced cancer specificity using sample volumes similar or lower than corresponding traditional ELISAs. For ovarian cancer, we show that this new approach when applied to ovarian cyst fluid samples provide results similar to the performance obtained with ctDNA determinations of a set of 17 driver mutations and greatly superior compared to corresponding conventional immunoassays. Based on our results, we predict that the nanoparticle-lectin concept will enable a new generation of simple, low-cost biomarker assays of highly improved cancer specificity. Such tools should ideally be evaluated together with determination of ctDNA to establish early detection schemes for cancers e.g. ovarian, pancreas, lung where the detection rate of early stage disease is presently unacceptably low.</div

Pandemic influenza A(H1N1 pdm09) vaccine induced high levels of influenza-specific IgG and IgM antibodies as analyzed by enzyme immunoassay and dual-mode multiplex microarray immunoassay methods

Influenza A viruses continue to circulate throughout the world as yearly epidemics or occasional pandemics. Influenza infections can be prevented by seasonal multivalent or monovalent pandemic vaccines. In the present study, we describe a novel multiplex microarray immunoassay (MAIA) for simultaneous measurement of virus-specific IgG and IgM antibodies using Pandemrix-vaccinated adult sera collected at day 0 and 28 and 180 days after vaccination as the study material. MAIA showed excellent correlation with a conventional enzyme immunoassay (EIA) in both IgG and IgM anti-influenza A antibodies and good correlation with hemagglutination inhibition (HI) test. Pandemrix vaccine induced 5-30 fold increases in anti-H1N1pdm09 influenza antibodies as measured by HI, EIA or MAIA. A clear increase in virus-specific IgG antibodies was found in 93-97% of vaccinees by MAIA and EIA. Virus-specific IgM antibodies were found in 90-92% of vaccinees by MAIA and EIA, respectively and IgM antibodies persisted for up to 6 months after vaccination in 55-62% of the vaccinees. Pandemic influenza vaccine induced strong anti-influenza A IgG and IgM responses that persisted several months after vaccination. MAIA was demonstrated to be an excellent method for simultaneous measurement of antiviral IgG and IgM antibodies against multiple virus antigens. Thus the method is well suitable for large scale epidemiological and vaccine immunity studies. (C) 2020 Elsevier Ltd. All rights reserved

Serological Array-in-Well Multiplex Assay Reveals a High Rate of Respiratory Virus Infections and Reinfections in Young Children

Serological assays are used to diagnose and characterize host immune responses against microbial pathogens. Microarray technologies facilitate high-throughput immunoassays of antibody detection against multiple pathogens simultaneously. To improve survey of influenza A virus (IAV), influenza B virus (IBV), respiratory syncytial virus (RSV), and adenovirus (AdV) antibody levels, we developed a microarray consisting of IAV H1N1, IAV H1N1pdm09 (vaccine), IAV H3N2, IBV Victoria, IBV Yamagata, RSV, AdV type 5 hexon protein, and control antigens printed on the bottom of a microtiter plate well. Bound IgG antibodies were detected with anti-human IgG-coated photon-upconverting nanoparticles and measured with a photoluminescence imager. The performance of the microarray immunoassay (MAIA) was evaluated with serum samples (n = 576) collected from children (n - 288) at 1 and 2 years of age and tested by standard enzyme immunoassays (EIAs) for antibodies to IAV vaccine and RSV. EIAs and MAIA showed substantial to almost perfect agreement (Cohen's kappa, 0.62 to 0.83). Applying MAIA, we found seroprevalences of 55% for IAV H1N1, 54% for IAV vaccine, 30% for IAV H3N2, 24% for IBV Victoria, 25% for IBV Yamagata, 38% for RSV, and 26% for AdV in 1-year-old children (n = 768). By the age of 2 years, IgG seropositivity rates (n = 714) increased to 74% for IAV H1N1, 71% for IAV vaccine, 49% for IAV H3N2, 47% for IBV Yamagata, 49% for IBV Victoria, 68% for RSV, and 58% for AdV. By analyzing increases in antibody levels not biased by vaccinations, we found a reinfection rate of 40% for RSV and 31% for AdV in children between 1 and 2 years of age.IMPORTANCE The multiplex immunoassay was successfully used to simultaneously detect antibodies against seven different viruses. The developed serological microarray is a new promising tool for diagnostic, epidemiological, and seroprevalence analyses of virus infections