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Mapix

Manufactured by Innopsys
Sourced in France, United States

Mapix is a versatile and high-performance microscope imaging system designed for a wide range of applications. It integrates advanced imaging capabilities to capture detailed and accurate images of samples.

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23 protocols using mapix

1

Biotinylated Viral Glycan Binding Assay

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Viruses were directly labeled with a biotin handle as previously described (Watanabe et al., 2014 (link)). Labeled viruses were diluted to 256 hemagglutinating units (HAU) in 1X PBS and applied directly to the slide surface for 1h. Following the initial incubation, arrays were washed, by dipping 3 times in 1X PBS and again 3 times in 1X PBS. Washed arrays were incubated with 2ug/mL streptavidin-AlexaFluor555 (LifeTechnologies) in 1X PBS, for 1h. Following detection, arrays were washed sequentially, by dipping, 3 times in 1X PBS, 3 times in 1X PBS and, finally, 3 times in deionized H2O. Washed arrays were dried by centrifugation and immediately scanned for AlexaFluor555 signal on an Innoscan 1100AL (Innopsys) confocal microarray scanner. Signal intensity from scanned arrays was collected using Mapix (Innopsys). Signal intensity was calculated for the mean signal intensity of 4 replicate spots for each printed glycan and graphed using Excel (Microsoft).
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2

Array CGH Analysis of Rare Genetic Disorders

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Array CGH was performed for all the components of the four families after DNA extraction (QIAmp DNA blood Maxi Kit, Qiagen, Hilden, Germany) from patients and relatives peripheral blood cells. CytoSure ISCA V2 4x180K platform with a backbone resolution of 1 probe/25 Kb and 1 probe/19 Kb in critical regions, human genome reference GRCh37/hg19 and sex matched normal human DNA pool (Kreatech, Amsterdam, Netherlands) as control were used. InnoScan 710 Microarray Scanner (Innopsys, Carbonne, France) and Mapix (Innopsys, Carbonne, France) were used to detect and analyze fluorescence levels, respectively. Results were interpreted using Cytosure Interpret Software (Oxford Gene Technology, Begbroke, United Kingdom). QC metrics: SD < 1.0 and DLR spread <0.3 were required. The significance of CNVs (copy number variants) was evaluated according to American College of Medical Genetics (ACMG) Joint Consensus (Riggs et al., 2012 (link)).
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3

Fluorescence Imaging of Labeled Antibodies

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Dried slides were imaged using an ImageXpress imaging system to detect fluorescently labeled secondary antibodies. The imager used an LED light engine (SemRock) centered at 532nm wavelength to excite fluorophore-conjugated secondary Ab. Mapix (version 7.2.1; Innopsys, Carbonne, France) was used to place a grid alignment file over the obtained images and extract the median foreground pixel intensities using the central 60% of each feature. Images were saved as TIFF files and extracted intensities saved as GenePix results files.
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4

Glycan Array Analysis of Recombinant HA

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Recombinant trimeric HA0 was expressed in HEK293S GnTI-/- (American Type Culture Collection), purified and analyzed on glycan array as previously described [35 (link)]. Briefly, soluble trimeric HA (50 μg mL-1) was pre-complexed with the anti-HIS mouse antibody (MA1-21315, Thermo Fisher Scientific, Waltham, MA) and the Alexa647-linked anti-mouse IgG (A-21235, Thermo Fisher Scientific) at 4:2:1 molar ratio for 15 min on ice in 50 μL PBST. This complex was incubated on the array surface in a humidified chamber for 60 min before washing and analysis using an Innoscan 1100AL microarray scanner (Innopsys, Chicago, IL). Fluorescent signal intensity was measured using Mapix (Innopsys) and mean intensity minus mean background of 4 replicate spots was calculated. A complete list of the glycans on the array is presented in S11 Fig.
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5

Gene Expression Analysis of Exposure

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The inverted TIFF images were processed with Mapix (Innopsys) to ascribe a value to the spot intensity, which was corrected for background intensity. The intensity data were normalized within and between arrays using the Bioconductor package limma using the control reference genes RPL8 and actin. To test for differential expression, the bayesian adjusted t-statistics from the linear models for Microarray data (limma) package was used on the combined dataset [28 (link), 29 ]. The combined dataset possesses the data originating from 8 biological replicates and the technical repetition was also included. Adjusted p-values were calculated using the method developed by Benjamin and Hochberg [30 ]. The differential expression was calculated by comparing the expression data from an exposed experiment with its corresponding control experiment. Thus, the exposed cage experiments were compared with the non exposed cage experiments from the same exposure time and the exposed field experiments were compared to the non-exposed field experiments with the corresponding exposure time.
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6

Comprehensive Angiogenesis and MMP Profiling in Tissue and Saliva

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Tissue and saliva samples from patients were used to measure the expression of ANG, angiopoietin-2 (ANG-2), bFGF, heparin-binding EGF (HB-EGF), HGF, LEP, PDGF-BB, PIGF, and VEGF by the Quantibody® Human Angiogenesis Array 1 (RayBiotech, USA) and MMP-1, MMP-2, MMP-3, MMP-8, MMP-9, MMP-10, MMP-13, TIMP-1, and TIMP-2 by the Quantibody® Human MMP Array 1 (RayBiotech, USA). The detection was performed according to the manufacturer's instructions. Briefly, supernatant from centrifuged tissue samples treated with cell lysate and centrifuged saliva samples were used. The glass slide was completely air dry and blocked by a blocking solution for 1 h. The tissue sample diluent 100 µL (500 µg/mL) and 80 µL of the diluted tissue lysate or saliva samples were separately added to the slides and incubated overnight at 4˚C. The samples diluted were decanted from the wells and washed with wash buffer at room temperature with gentle rocking. The detection antibody cocktail was added to each well and incubated at room temperature, followed by a washing step. Afterward, Cy3 equivalent dye-conjugated streptavidin was added and incubated in a dark room, followed by a washing step. Fluorescence signals were obtained by an InnoScan 300 Microarray Scanner (Innopsys, France) at 532 nm wavelength and 10 µm resolution. Data were analyzed using Mapix (Innopsys, France) analysis software.
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7

Quantitative Protein Array Analysis

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Cell lysates were prepared and analyzed as previously described (Teo et al., 2018 (link)). Briefly, medium was removed from the plates and cells were rinsed twice with PBS. Cells were lysed on ice for 20 minutes with occasional shaking every 5 minutes. Cell lysates were collected with a scraper and clarified by centrifugation at 18,000 rcf. for 10 min at 4°C. Clarified supernatants in biological triplicate were adjusted to 2 mg/mL concentration and printed onto nitrocellulose-coated slides (Grace Bio-Labs) in a dilution series (four serial 2-fold dilutions) in technical triplicate using an Aushon 2470 arrayer (Aushon Biosystems). Slides were blocked, probed with validated primary antibodies and detected with DyLight 800-conjugated secondary antibodies (New England BioLabs). Slides were read using an InnoScan 710-IR scanner (Innopsys) and quantified using Mapix (Innopsys). Relative fluorescence intensities were normalized to respective FastGreen-stained spots (total protein), and data were computationally analyzed as previously described (Byron, 2017 (link); Teo et al., 2018 (link)).
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8

Glycan Array Analysis of Antibody Binding to Influenza NA Antigen

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The Sh2 N9 NA tetramer with ectodomain plus stalk and His6-tag was diluted to 20 pg/mL and incubated with antibody IgG at a molar ratio of one NA protomer to four antibody IgGs for 1 hour at room temperature (~ 22 °C) in 100 mM imidazole-malate pH 6.15, 150 mM NaCl, 10 mM CaCl2, and 0.02% NaN3. The mixtures were then applied to glycan arrays (see Data S1 for glycan list) (Peng et al., 2017 (link)) for 1 hour. Then each array was washed to remove the NA solution by dipping 3 times with 1× PBS + 0.05% Tween, pH 7.4 at room temperature. Following washing, 250 μL of a pre-complexed solution of biotinylated Erythrina cristagalli lectin (ECA) (10 μg/mL, VectorLabs) + Streptavidin-AlexaFluor555 (2 μg/mL, Thermo Fisher Scientific) was applied directly to the array surface and incubated for 2 hours. Following incubation, ECA-Streptavidin solution was removed by dipping 3 times with 1× PBS + 0.05% Tween and, subsequently, by dipping 3 times in 1× PBS and then 3 times in distilled H2O. Washed slides were dried by centrifugation and scanned on an Innoscan1100AL (Innopsys) confocal slide scanner for 532 emission. Image data were stored as a TIFF image and signal data were collected using Mapix (Innopsys) imaging software. Collected signal data were processed to determine the averaged (mean signal minus mean background) values of 4 replicate spots on the array for each unique printed glycan.
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9

Fluorescence Image Analysis Protocol

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Fluorescence scanning was performed on the Innoscan 1100 AL (Innopsys, Carbonne, France) at 647 nm, gain 3, laser power low, and a resolution of 1 μm/pixel. The resulting 16-bit grayscale images were analyzed using Mapix (Innopsys, Carbonne, France). The median gray scale value of all pixels in a spot was subtracted by the median gray scale value of at least 5 background control areas (spot size of 80–90 µm each), located in the vicinity of the spot of interest.
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10

Quantitative Microarray Image Analysis

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To enable quantification of the dye concentration indicative of the antibody concentration in all spots of the array, the intensity must be determined in each spot. For this purpose, microarray slides were printed with a distinct pattern of 18 × 18 dots (150 μm diameter, 280 μm spacing). The slides were imaged using both the TinyArray imager and the commercial ArrayCam system (Grace Bio-Labs, Bend, OR). As the size and distances of the spots were known, the background-subtracted median spot fluorescence intensity was measured and quantified using Mapix (Innopsys) as previously described.25–27 (link) Mean fluorescence intensity (MFI) of the four replicate spots for each antigen was utilized for antigen heat map generation and data analysis.
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