microcaster hand-held microarrayer system  (Schleicher Inc)

Journal: Nucleic Acids Research

Article Title: Functional studies on transfected cell microarray analysed by linear regression modelling

doi: 10.1093/nar/gkn428

Figure Lengend Snippet: Effects of sucrose and gelatine concentrations on spot integrity and transfection efficiency. ( A ) Array printed with pDsRed (50 ng/µl) in a printing solution with different gelatine and sucrose concentrations. Scanning image of the whole array and magnifications of specific spots. ( B ) Array printed with pEGFP (50 ng/µl) in a printing solution with 25 mM sucrose and four different concentrations of gelatine. Top: Box plot of the fluorescence intensities in each spot ( n = 32–34). Bottom : Scanning image showing squares of seven times five spots for the four gelatine concentrations. From left to right: 0.01, 0.05, 0.1 and 0.2% gelatine. The DNA-lipid-gelatine-sucrose solutions were printed manually with a 10 µl pipette tip (A) or by MicroCaster TM manual arrayer system (B).

Article Snippet: We also used the hand-held microarrayer MicroCaster TM from Schleicher and Schuell, which consists of an arrayer tool containing eight pins and a slide holder with an indexing system to guide the spotting of up to 768 spots, each with a diameter of about 500 μm (about 6 nl sample per spot).

Techniques: Transfection, Fluorescence, Transferring

Journal: Nucleic Acids Research

Article Title: Functional studies on transfected cell microarray analysed by linear regression modelling

doi: 10.1093/nar/gkn428

Figure Lengend Snippet: Evaluation of fluorescence intensities in each spot. ( A ) Density plots of pixel intensities from six spots printed with pEGFP (green lines). Mean (black lines) and median (red lines) of the pixel intensities in each spot. The scanning image shows the EGFP fluorescence intensity signal (green) in the six spots. The pixel intensities in each spot were acquired using the ‘Example save pixel values’ in the Report menu in GenePix software. This resulted in a text-file with 7884 pixel values for each spot. Plotting was done using the R software. ( B ) Spot-to-spot variability in 26 spots printed with pEGFP and pDsRed. EGFP, DsRed and ratio (EGFP normalized to DsRed) fluorescence intensity signal (mean of the pixel intensities) normalized to the mean of the signal from all the spots. The arrays were printed using MicroCaster TM .

Article Snippet: We also used the hand-held microarrayer MicroCaster TM from Schleicher and Schuell, which consists of an arrayer tool containing eight pins and a slide holder with an indexing system to guide the spotting of up to 768 spots, each with a diameter of about 500 μm (about 6 nl sample per spot).

Techniques: Fluorescence, Software

Journal: Nucleic Acids Research

Article Title: Functional studies on transfected cell microarray analysed by linear regression modelling

doi: 10.1093/nar/gkn428

Figure Lengend Snippet: Linear regression modelling without a treatment effect. Cells were cotransfected with pEGFP (30 ng/µl), pDsRed (30 ng/µl) and different concentrations of siEGFP (0–30 ng/µl). ( A ) Left diagram indicates the placement of the cell clusters for eight conditions. pEGFP, pDsRed and 0, 2.5, 5, 10, 15 or 30 ng/µl siEGFP. The non-fluorescent reporter pCRE-Luc (60 ng/µl) as a control of the background fluorescence in the spots (negctrl). pEGFP, pDsRed and 30 ng/µl siCAT as a siRNA control (ctrl). Middle diagram: Scanning image for EGFP-detection. Right diagram: Scanning image for DsRed-detection. ( B ) Illustration of the different terms in the linear regression model in Equation . The ratio fluorescence signal intensities (EGFP intensities normalized to DsRed intensities) from each spot (log-transformed and normalized to negative control spot intensities) in two experimental replicates (28–35 technical replicates after removing flagged spots) were used to fit the model. (a) The overall effect µ . (b) The overall effect µ and the experimental replicate effects e and e 2 . (c) The overall effect µ and the condition effects c through c 8 . (d) Combining µ, e i and c k . (e) Addition of the interaction effect term (ec) ik to the terms in (d). The observed data from each spot are shown together with the fitted model in (d) and (e) for experimental replicate 1 (black circles) and experimental replicate 2 (blue triangles). ( C ) Estimated effects of the different conditions are shown based on the EGFP fluorescence intensity data (green, left) or the EGFP fluorescence intensity data normalized to DsRed control plasmid intensities (ratio, right). The intensities are shown relative to 0 ng/µl siRNA, error bars are 95% CI. The arrays were printed using MicroCaster TM .

Article Snippet: We also used the hand-held microarrayer MicroCaster TM from Schleicher and Schuell, which consists of an arrayer tool containing eight pins and a slide holder with an indexing system to guide the spotting of up to 768 spots, each with a diameter of about 500 μm (about 6 nl sample per spot).

Techniques: Control, Fluorescence, Transformation Assay, Negative Control, Plasmid Preparation